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WO2024006842A2 - Épitopes de lymphocytes t de bordetella, mégapools et utilisations associées - Google Patents

Épitopes de lymphocytes t de bordetella, mégapools et utilisations associées Download PDF

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Publication number
WO2024006842A2
WO2024006842A2 PCT/US2023/069274 US2023069274W WO2024006842A2 WO 2024006842 A2 WO2024006842 A2 WO 2024006842A2 US 2023069274 W US2023069274 W US 2023069274W WO 2024006842 A2 WO2024006842 A2 WO 2024006842A2
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WO
WIPO (PCT)
Prior art keywords
pertussis
peptides
amino acid
set forth
bordetella
Prior art date
Application number
PCT/US2023/069274
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English (en)
Other versions
WO2024006842A3 (fr
Inventor
Alessandro Sette
Ricardo Da Silva ANTUNES
Original Assignee
La Jolla Institute For Immunology
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Filing date
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Application filed by La Jolla Institute For Immunology filed Critical La Jolla Institute For Immunology
Publication of WO2024006842A2 publication Critical patent/WO2024006842A2/fr
Publication of WO2024006842A3 publication Critical patent/WO2024006842A3/fr

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Classifications

    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • A61K39/02—Bacterial antigens
    • A61K39/099—Bordetella
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
    • A61P31/04—Antibacterial agents
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • C07K14/195—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from bacteria
    • C07K14/235—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from bacteria from Bordetella (G)
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • A61K2039/51—Medicinal preparations containing antigens or antibodies comprising whole cells, viruses or DNA/RNA
    • A61K2039/52—Bacterial cells; Fungal cells; Protozoal cells
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • A61K2039/57—Medicinal preparations containing antigens or antibodies characterised by the type of response, e.g. Th1, Th2
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • A61K2039/70—Multivalent vaccine

Definitions

  • the present invention relates in general to the field of proteins and peptides that are T cell epitopes and/or antigens for Bordetella, including epitopes and antigens from B. pertussis, and more particularly, to compositions and methods for the prevention, treatment, diagnosis, kits, and uses of such T cell epitopes and antigens, including megapools, for use in detecting and characterizing B. pertussis specific responses in infection and following vaccination.
  • the present invention includes a composition comprising: one or more peptides or proteins, comprising, consisting of, or consisting essentially of an amino acid sequence selected from the sequences set forth in any one of Tables 1-20 (SEQ ID NOS: 1 to 2598), or a subsequence, portion, homologue, variant or derivative thereof; a fusion protein comprising one or more amino acid sequences selected from any one of those sequences set forth in Tables 1-20; a pool of 2 or more or more peptides comprising, consisting of, or consisting essentially of amino acid sequences selected from any one of those sequences set forth in Tables 1-20; or a polynucleotide that encodes one or more peptides or proteins, comprising, consisting of, or consisting essentially of an amino acid sequence selected from any one of those sequences set forth in Tables 1-20, or a subsequence, portion, homologue, variant or derivative thereof.
  • the one or more peptides or proteins comprises, or wherein the fusion protein comprises 2 or more or more amino acid sequences selected from any one of those sequences set forth in Tables 1-20, or a subsequence, portion, homologue, variant or derivative thereof.
  • the amino acid sequence is selected from a Bordetella T cell epitope selected from any one of those sequences set forth in Tables 1-20.
  • the composition comprises one or more B.
  • the peptide or protein comprises a Bordetella T cell epitope.
  • the one or more peptides or proteins comprises a Bordetella CD8+ or CD4+ T cell epitope.
  • the Bordetella is B. pertussis and the B.
  • pertussis T cell epitope is not conserved in another Bordetella.
  • the Bordetella is B. pertussis and the B. pertussis T cell epitope is conserved in another Bordetella.
  • the one or more peptides or proteins has a length from about 9-15, 15-20, 20-25, 25-30, 30-40, 40-50, 50-75 or 75- 100 amino acids.
  • the one or more peptides or proteins elicits, stimulates, induces, promotes, increases or enhances a T cell response to a Bordetella.
  • the one or more peptides or proteins that elicits, stimulates, induces, promotes, increases or enhances the T cell response to the Bordetella is a Bordetella protein or peptide, or a variant, homologue, derivative or subsequence thereof.
  • the composition further comprises formulating the one or more peptides or proteins into an immunogenic formulation with an adjuvant.
  • the adjuvant is selected from the group consisting of adjuvant is selected from the group consisting of alum, aluminum hydroxide, aluminum phosphate, calcium phosphate hydroxide, cytosine-guanosine oligonucleotide (CpG-ODN) sequence, granulocyte macrophage colony stimulating factor (GM-CSF), monophosphoryl lipid A (MPL), poly(I:C), MF59, Quil A, N-acetyl muramyl-L-alanyl-D-isoglutamine (MDP), FIA, montanide, poly (DL- lactide-coglycolide), squalene, virosome, AS03, ASO4, IL-1, IL-2, IL-3, IL-4, IL-5, IL-6, IL-7, IL-8, IL- 10, IL-12, IL-15, IL-17, IL-18, STING, CD40L, pathogen-associated molecular patterns (PAMPs), pathogen
  • the composition further comprises a modulator of immune response.
  • the modulator of immune response is a modulator of the innate immune response.
  • the modulator is Interleukin-6 (IL-6), Interferon-gamma (IFN-y), Transforming growth factor beta (TGF-P), or Interleukin- 10 (IL-10), or an agonist or antagonist thereof.
  • the present invention includes a composition comprising monomers or multimers of: peptides or proteins comprising, consisting of, or consisting essentially of: one or more amino acid sequences selected from any one of those sequences set forth in Tables 1-20, concatemers, subsequences, portions, homologues, variants or derivatives thereof; a fusion protein comprising one or more amino acid sequences selected from any one of those sequences set forth in Tables 1-20; or a polynucleotide that encodes one or more peptides or proteins, comprising, consisting of, or consisting essentially of an amino acid sequence selected from any one of those sequences set forth in Tables 1-20, or a subsequence, portion, homologue, variant or derivative thereof.
  • the present invention includes a composition comprising one or more peptide-major histocompatibility complex (MHC) monomers or multimers, wherein the peptide-MHC monomer or multimer comprises a peptide comprising, consisting of, or consisting essentially of an amino acid sequence selected from any one of those sequences set forth in Tables 1-20, in a groove of the MHC monomer or multimer.
  • MHC peptide-major histocompatibility complex
  • the present invention includes a composition comprising: one or more peptides or proteins comprising, consisting of, or consisting essentially of an amino acid sequence selected from any one of those sequences set forth in Tables 1-20, or a subsequence, portion, homologue, variant or derivative thereof; a fusion protein comprising one or more amino acid sequences selected from any one of those sequences set forth in Tables 1-20; a pool of 2 or more peptides selected from any one of those sequences set forth in Tables 1-20; a polynucleotide that encodes one or more peptides or proteins, comprising, consisting of, or consisting essentially of an amino acid sequence selected from any one of those sequences set forth in Tables 1-20, or a subsequence, portion, homologue, variant or derivative thereof.
  • the one or more peptides or proteins comprises, or wherein the fusion protein comprises, 2 or more amino acid sequences selected from any one of those sequences set forth in Tables 1-20, or a subsequence, portion, homologue, variant or derivative thereof.
  • the protein or peptide comprises a B. pertussis T cell epitope.
  • the one or more peptides or proteins comprises a B. pertussis CD8+ or CD4+ T cell epitope.
  • the B. pertussis T cell epitope is not conserved in another Bordetella.
  • the B. pertussis T cell epitope is conserved in another Bordetella.
  • the one or more peptides or proteins has a length from about 9-15, 15-20, 20-25, 25-30, 30-40, 40-50, 50-75 or 75-100 amino acids.
  • the one or more peptides or proteins elicits, stimulates, induces, promotes, increases or enhances a T cell response to B. pertussis.
  • the one or more peptides or proteins that elicits, stimulates, induces, promotes, increases or enhances the T cell response to B. pertussis is a B. pertussis protein or peptide, or a variant, homologue, derivative or subsequence thereof.
  • the composition further comprises formulating the one or more peptides or proteins into an immunogenic formulation with an adjuvant.
  • the adjuvant is selected from the group consisting of adjuvant is selected from the group consisting of alum, aluminum hydroxide, aluminum phosphate, calcium phosphate hydroxide, cytosine-guanosine oligonucleotide (CpG-ODN) sequence, granulocyte macrophage colony stimulating factor (GM-CSF), monophosphoryl lipid A (MPL), poly(I:C), MF59, Quil A, N-acetyl muramyl-L-alanyl-D-isoglutamine (MDP), FIA, montanide, poly (DL-lactide-coglycolide), squalene, virosome, AS03, ASO4, IL-1, IL-2, IL- 3, IL-4, IL-5, IL-6, IL-7, IL-8, IL-10, IL-12,
  • the composition further comprises a modulator of immune response.
  • the modulator of immune response is a modulator of the innate immune response.
  • the modulator is Interleukin-6 (IL-6), Interferon-gamma (IFN- y), Transforming growth factor beta (TGF-P), or Interleukin- 10 (IL- 10), or an agonist or antagonist thereof.
  • the present invention includes a composition comprising monomers or multimers of: one or more peptides or proteins comprising, consisting of, or consisting essentially of: one or more B.
  • pertussis amino acid sequences selected from any one of those sequences set forth in Tables 1- 20, concatemers, subsequences, portions, homologues, variants or derivatives thereof; a fusion protein comprising one or more amino acid sequences selected from any one of those sequences set forth in Tables 1-20; or a polynucleotide that encodes one or more peptides or proteins, comprising, consisting of, or consisting essentially of an amino acid sequence selected from any one of those sequences set forth in Tables 1-20, or a subsequence, portion, homologue, variant or derivative thereof.
  • the present invention includes a composition comprising one or more peptide-major histocompatibility complex (MHC) monomers or multimers, wherein the peptide-MHC monomer or multimer comprises a peptide comprising, consisting of, or consisting essentially of an amino acid sequence selected from any one of those sequences set forth in Tables 1-20, in a groove of the (MHC) monomer or multimer.
  • MHC peptide-major histocompatibility complex
  • the present invention includes a method for detecting the presence of: (i) a Bordetella or (ii) an immune response relevant to Bordetella infections, vaccines or therapies, including T cells responsive to one or more Bordetella peptides, comprising: providing one or more proteins or peptides for detection of an amount or a relative amount of, and/or the activity of, and/or the state of antigen-specific T-cells; contacting a biological sample suspected of having Bordetella-specific T-cells to one or more proteins or peptides for detection; and detecting an amount or a relative amount of, and/or the activity of, and/or the state of antigen-specific T-cells in the biological sample, wherein the one or more proteins or peptides for detection comprise one or more amino acid sequences set forth in any one of Tables 1-20, or comprise a pool of 2 or more or more amino acid sequences set forth in any one of Tables 1-20.
  • detecting the amount or a relative amount of, and/or activity of antigen-specific T-cells comprises one or more steps of identification or detection of the antigen-specific T-cells and measuring the amount of the antigen-specific T-cells.
  • the one or more peptides or proteins comprises 2 or more amino acid sequences selected from any one of Tables 1-20.
  • the detecting the amount or a relative amount of, and/or activity of antigen-specific T-cells comprises indirect detection and/or direct detection.
  • the method of detecting an immune response relevant to the Bordetella comprises the following steps: providing an MHC monomer or an MHC multimer; contacting a population T-cells to the MHC monomer or MHC multimer; and measuring the number, activity or state of T-cells specific for the MHC monomer or MHC multimer.
  • the MHC monomer or MHC multimer comprises a protein or peptide of the Bordetella.
  • the protein or peptide comprises a CD8+ or CD4+ T cell epitope.
  • the T cell epitope is not conserved in another Bordetella.
  • the T cell epitope is conserved in another Bordetella.
  • the protein or peptide has a length from about 9-15, 15-20, 20-25, 25-30, 30-40, 40-50, 50-75 or 75-100 amino acids.
  • the proteins or peptides comprise 2 or more amino acid sequences selected from any one of those sequences set forth in Tables 1-20, or a subsequence, portion, homologue, variant or derivative thereof.
  • the method further comprises detecting the presence or amount of the one or more peptides in a biological sample, or a response thereto, which is diagnostic of a Bordetella infection.
  • the detecting an amount or a relative amount of, and/or the activity of, and/or the state of antigen-specific T-cells in the biological sample comprises measuring one or more of a cytokine or lymphokine secretion assay, T cell proliferation, immunoprecipitation, immunoassay, ELISA, radioimmunoassay, immunofluorescence assay, Western Blot, FACS analysis, a competitive immunoassay, a noncompetitive immunoassay, a homogeneous immunoassay a heterogeneous immunoassay, a bioassay, a reporter assay, a luciferase assay, a microarray, a surface plasmon resonance detector, a florescence resonance energy transfer, immunocytochemistry, or a cell mediated assay, or a cytokine proliferation assay.
  • the method further comprises administering a treatment comprising the composition of one or more proteins, peptides or multimers to the subject from which the
  • the present invention includes a method for detecting the presence of: (i) B. pertussis or (ii) an immune response relevant to B. pertussis infections, vaccines or therapies, including T cells responsive to one or more B. pertussis peptides, comprising: providing one or more proteins or peptides for detection of an amount or a relative amount of, and/or the activity of, and/or the state of antigen-specific T-cells; contacting a biological sample suspected of having B.
  • detecting the amount or a relative amount of, and/or activity of antigen-specific T-cells comprises one or more steps of identification or detection of the antigen-specific T-cells and measuring the amount of the antigen-specific T-cells.
  • the one or more peptides or proteins comprises 2 or more amino acid sequences selected from any one of those sequences set forth in Tables 1-20.
  • detecting the amount or a relative amount of, and/or activity of antigen-specific T-cells comprises indirect detection and/or direct detection.
  • detecting an immune response relevant to B. pertussis comprises the following steps: providing an MHC monomer or an MHC multimer; contacting a population T-cells to the MHC monomer or MHC multimer; and measuring the number, activity or state of T-cells specific for the MHC monomer or MHC multimer.
  • the MHC monomer or MHC multimer comprises a protein or peptide of B. pertussis.
  • the protein or peptide comprises a B. pertussis CD8+ or CD4+ T cell epitope.
  • the B. pertussis T cell epitope is not conserved in another Bordetella.
  • the B. pertussis T cell epitope is conserved in another Bordetella.
  • the protein or peptide has a length from about 9-15, 15-20, 20-25, 25-30, 30-40, 40-50, 50-75 or 75-100 amino acids.
  • the proteins or peptides comprise 2 or more amino acid sequences selected from any one of those sequences set forth in Tables 1-20, or a subsequence, portion, homologue, variant or derivative thereof.
  • the method further comprises detecting the presence or amount of the one or more peptides in a biological sample, or a response thereto, which is diagnostic of a B. pertussis infection.
  • detecting an amount or a relative amount of, and/or the activity of, and/or the state of antigen-specific T- cells in the biological sample comprises measuring one or more of a cytokine or lymphokine secretion assay, T cell proliferation, immunoprecipitation, immunoassay, ELISA, radioimmunoassay, immunofluorescence assay, Western Blot, FACS analysis, a competitive immunoassay, a noncompetitive immunoassay, a homogeneous immunoassay a heterogeneous immunoassay, a bioassay, a reporter assay, a luciferase assay, a microarray, a surface plasmon resonance detector, a florescence resonance energy transfer, immunocytochemistry, or a cell mediated assay
  • the method further comprises administering a treatment comprising the composition of one or more proteins, peptides or multimers to the subject from which the biological sample was drawn that increases the amount or relative amount of, and/or activity of the antigen-specific T-cells.
  • the present invention includes a method detecting a Bordetella infection or exposure in a subject, the method comprising, consisting of, or consisting essentially of: contacting a biological sample from a subject with a composition of composition of one or more proteins, peptides or multimers; and determining if the composition elicits an immune response from the contacted cells, wherein the presence of an immune response indicates that the subject has been exposed to or infected with Bordetella.
  • the sample comprises T cells.
  • the response comprises inducing, increasing, promoting or stimulating axtii-Bordetella activity of T cells.
  • the T cells are CD8+ or CD4+ T cells.
  • the method comprises determining whether the subject has been infected by or exposed to the Bordetella more than once by determining if the subject elicits a secondary T cell immune response profile that is different from a primary T cell immune response profile.
  • the method further comprises diagnosing a Bordetella infection or exposure in a subject, the method comprising contacting a biological sample from a subject with a composition of composition of one or more proteins, peptides or multimers, and determining if the composition elicits a T cell immune response, wherein the T cell immune response identifies that the subject has been infected with or exposed to a Bordetella.
  • the method is conducted three or more days following the date of suspected infection by or exposure to a Bordetella.
  • the present invention includes a method detecting B. pertussis infection or exposure in a subject, the method comprising, consisting of, or consisting essentially of: contacting a biological sample from a subject with a composition of composition of one or more proteins, peptides or multimers; and determining if the composition elicits an immune response from the contacted cells, wherein the presence of an immune response indicates that the subject has been exposed to or infected with B. pertussis.
  • the sample comprises T cells.
  • the response comprises inducing, increasing, promoting or stimulating anti -B. pertussis activity of T cells.
  • the T cells are CD8+ or CD4+ T cells.
  • the method comprises determining whether the subject has been infected by or exposed to B. pertussis more than once by determining if the subject elicits a secondary T cell immune response profde that is different from a primary T cell immune response profde.
  • the method further comprises diagnosing a B. pertussis infection or exposure in a subject, the method comprising contacting a biological sample from a subject with a composition of one or more proteins, peptides or multimers; and determining if the composition elicits a T cell immune response, wherein the T cell immune response identifies that the subject has been infected with or exposed to B. pertussis.
  • the method is conducted three or more days following the date of suspected infection by or exposure to a Bordetella.
  • the present invention includes a kit for the detection of Bordetella or an immune response to Bordetella in a subject comprising, consisting of or consisting essentially of: one or more T cells that specifically detect the presence of: one or more amino acid sequences selected from any one of those sequences set forth in Tables 1-20, or a subsequence, portion, homologue, variant or derivative thereof; or a fusion protein comprising one or more amino acid sequences selected from any one of those sequences set forth in Tables 1-20; or a pool of 2 or more or more peptides selected from the amino acid sequences set forth in any one of Tables 1-20.
  • the one or more amino acid sequences are selected from a Bordetella T cell epitope set forth in any one of Tables 1-20.
  • the composition comprises: one or more amino acid sequences selected from any one of those sequences set forth in Tables 1-20, or a subsequence, portion, homologue, variant or derivative thereof; a fusion protein comprising one or more amino acid sequences selected from any one of those sequences set forth in Tables 1-20; or a pool of 2 or more peptides selected from the amino acid sequences set forth in those sequences set forth in any one of Tables 1-20.
  • the amino acid sequence comprises a Bordetella CD8+ or CD4+ T cell epitope.
  • the T cell epitope is not conserved in another Bordetella. In another aspect, the T cell epitope is conserved in another Bordetella. In another aspect, the fusion protein has a length from about 9-15, 15-20, 20-25, 25-30, 30-40, 40-50, 50-75 or 75-100 amino acids.
  • the kit includes instruction for a diagnostic method, a process, a composition, a product, a service or component part thereof for the detection of: (i) Bordetella or (ii) an immune response relevant to Bordetella infections, vaccines or therapies, including T cells responsive to Bordetella.
  • the kit includes reagents for detecting an amount or a relative amount of, and/or the activity of, and/or the state of antigen-specific T-cells in the biological sample comprises measuring one or more of a cytokine or lymphokine secretion assay, T cell proliferation, immunoprecipitation, immunoassay, ELISA, radioimmunoassay, immunofluorescence assay, Western Blot, FACS analysis, a competitive immunoassay, a noncompetitive immunoassay, a homogeneous immunoassay a heterogeneous immunoassay, a bioassay, a reporter assay, a luciferase assay, a microarray, a surface plasmon resonance detector, a florescence resonance energy transfer, immunocytochemistry, or a cell mediated assay, or a cytokine proliferation assay.
  • the kit includes reagents for determining a Human Leukocyte Antigen (HLA) profile of a subject, and selecting peptides that are presented by the HLA profile of the subject for detecting an immune response to Bordetella.
  • HLA Human Leukocyte Antigen
  • the present invention includes a kit for the detection of B. pertussis or an immune response to B.
  • pertussis in a subject comprising, consisting of or consisting essentially of: one or more T cells that specifically detect the presence of: one or more amino acid sequences selected from any one of those sequences set forth in Tables 1-20, or a subsequence, portion, homologue, variant or derivative thereof; a fusion protein comprising one or more amino acid sequences selected from any one of those sequences set forth in Tables 1-20; or a pool of 2 or more peptides selected from the amino acid sequences set forth in those sequences set forth in any one of Tables 1-20.
  • the amino acid sequence comprises a B. pertussis CD8+ or CD4+ T cell epitope.
  • the B. pertussis CD8+ or CD4+ T cell epitope comprises a B. pertussis CD8+ or CD4+ T cell epitope.
  • kits includes instruction for a diagnostic method, a process, a composition, a product, a service or component part thereof for the detection of: (i) B. pertussis or (ii) an immune response relevant to B. pertussis infections, vaccines or therapies, including T cells responsive to B. pertussis.
  • the kit includes reagents for detecting an amount or a relative amount of, and/or the activity of, and/or the state of antigen-specific T- cells in the biological sample comprises measuring one or more of a cytokine or lymphokine secretion assay, T cell proliferation, immunoprecipitation, immunoassay, ELISA, radioimmunoassay, immunofluorescence assay, Western Blot, FACS analysis, a competitive immunoassay, a noncompetitive immunoassay, a homogeneous immunoassay a heterogeneous immunoassay, a bioassay, a reporter assay, a luciferase assay, a microarray, a surface plasmon resonance detector, a florescence resonance energy transfer, immunocytochemistry, or a cell mediated assay, or a cytokine proliferation assay.
  • the kit includes reagents for determining a Human Leukocyte Antigen (HLA) profile of a
  • the present invention includes a method of stimulating, inducing, promoting, increasing, or enhancing an immune response against a Bordetella in a subject, comprising: administering a composition of one or more proteins, peptides, multimers or a polynucleotide that expresses the protein, peptide or multimers, in an amount sufficient to stimulate, induce, promote, increase, or enhance an immune response against the Bordetella in the subject.
  • the immune response provides the subject with protection against a Bordetella infection or pathology, or one or more physiological conditions, disorders, illnesses, diseases or symptoms caused by or associated with Bordetella infection or pathology.
  • the immune response is specific to: one or more B. pertussis peptides selected from the amino acid sequences set forth in any one of Tables 1-20, or a subsequence, portion, homologue, variant or derivative thereof.
  • the present invention includes a method of stimulating, inducing, promoting, increasing, or enhancing an immune response against B. pertussis in a subject, comprising: administering a composition of proteins, peptides, multimers or a polynucleotide that expresses the protein, peptide or multimers, in an amount sufficient to stimulate, induce, promote, increase, or enhance an immune response against B. pertussis in the subject.
  • the immune response provides the subject with protection against a B. pertussis infection or pathology, or one or more physiological conditions, disorders, illnesses, diseases or symptoms caused by or associated with B. pertussis infection or pathology.
  • the immune response is specific to: one or more B. pertussis peptides selected from the amino acid sequences set forth in those sequences set forth in any one of Tables 1-20, or a subsequence, portion, homologue, variant or derivative thereof.
  • the present invention includes a method of stimulating, inducing, promoting, increasing, or enhancing an immune response against B. pertussis in a subject, comprising: administering to a subject an amount of a protein or peptide comprising, consisting of or consisting essentially of an amino acid sequence of a B. pertussis protein or peptide, or a variant, homologue, derivative or subsequence thereof, wherein the protein or peptide comprises at least two peptides selected from the amino acid sequences set forth in any one of Tables 1-20 or a subsequence, portion, homologue, variant or derivative thereof, in an amount sufficient to prevent, stimulate, induce, promote, increase, immunize against, or enhance an immune response against B. pertussis in the subject.
  • the immune response provides the subject with protection against B. pertussis infection or pathology, or one or more physiological conditions, disorders, illnesses, diseases or symptoms caused by or associated with B. pertussis infection or pathology.
  • the present invention includes a method of treating, preventing, or immunizing a subject against B. pertussis infection, comprising administering to a subject an amount of a protein or peptide comprising, consisting of, or consisting essentially of an amino acid sequence of a Bordetella protein or peptide, or a variant, homologue, derivative or subsequence thereof, wherein the protein or peptide comprises at least two amino acid sequences selected from any one of Tables 1-20 or a subsequence, portion, homologue, variant or derivative thereof, in an amount sufficient to treat, prevent, or immunize the subject for B.
  • the protein or peptide comprises or consists of a Bordetella T cell epitope that elicits, stimulates, induces, promotes, increases, or enhances an anti-5. pertussis T cell immune response.
  • the one or more amino acid sequences are selected from any one of those sequences set forth in Tables 1-20, or a subsequence, portion, homologue, variant or derivative thereof; a fusion protein comprising one or more amino acid sequences selected from any one of those sequences set forth in Tables 1-20; or a pool of 2 or more peptides selected from the amino acid sequences set forth in those sequences set forth in any one of Tables 1-20.
  • the anti -B is selected from any one of those sequences set forth in Tables 1-20, or a subsequence, portion, homologue, variant or derivative thereof.
  • the pertussis T cell response is a CD8+, a CD4+ T cell response, or both.
  • the T cell epitope is conserved across two or more clinical isolates of B. pertussis or two or more circulating forms of B. pertussis.
  • the B. pertussis infection is an acute infection.
  • the subject is a mammal or a human.
  • the method reduces B. pertussis bacterial titer, increases or stimulates B. pertussis bacterial clearance, reduces or inhibits B. pertussis bacterial proliferation, reduces or inhibits increases in B. pertussis bacterial titer or B. pertussis bacterial proliferation, reduces the amount of a B.
  • the method reduces one or more adverse physiological conditions, disorders, illness, diseases, symptoms or complications caused by or associated with B. pertussis infection or pathology. In another aspect, the method improves one or more adverse physiological conditions, disorders, illness, diseases, symptoms or complications caused by or associated with B. pertussis infection or pathology.
  • the symptom is fever or chills, cough, shortness of breath or difficulty breathing, fatigue, muscle or body aches, headache, new loss of taste or smell, sore throat, congestion or runny nose, nausea or vomiting, or diarrhea.
  • the method reduces or inhibits susceptibility to B. pertussis infection or pathology.
  • the protein or peptide, or a subsequence, portion, homologue, variant or derivative thereof is administered prior to, substantially contemporaneously with or following exposure to or infection of the subject with B. pertussis.
  • a plurality of B. pertussis T cell epitopes are administered prior to, substantially contemporaneously with or following exposure to or infection of the subject with B. pertussis.
  • the protein or peptide, or a subsequence, portion, homologue, variant or derivative thereof is administered within 2-72 hours, 2-48 hours, 4-24 hours, 4-18 hours, or 6-12 hours after a symptom of B. pertussis infection or exposure develops.
  • the protein or peptide, or a subsequence, portion, homologue, variant or derivative thereof is administered prior to exposure to or infection of the subject with B. pertussis.
  • the method further comprises administering a modulator of immune response prior to, substantially contemporaneously with or following the administration to the subject of an amount of a protein or peptide.
  • the modulator of immune response is a modulator of the innate immune response.
  • the modulator is IL-6, IFN-y, TGF-P, or IL- 10, or an agonist or antagonist thereof.
  • the present invention includes a method of treating, preventing, or immunizing a subject against B. pertussis infection, comprising administering to a subject the composition of one or more proteins, peptides or multimers in an amount sufficient to treat, prevent, or immunize the subject for B. pertussis infection.
  • the B. pertussis infection is an acute infection.
  • the method reduces B. pertussis bacterial titer, increases or stimulates B. pertussis bacterial clearance, reduces or inhibits B. pertussis bacterial proliferation, reduces or inhibits increases in B. pertussis bacterial titer or B. pertussis bacterial proliferation, reduces the amount of a B.
  • the method reduces one or more adverse physiological conditions, disorders, illness, diseases, symptoms or complications caused by or associated with B. pertussis infection or pathology. In another aspect, the method improves one or more adverse physiological conditions, disorders, illness, diseases, symptoms or complications caused by or associated with B. pertussis infection or pathology.
  • the symptom is fever or chills, cough, shortness of breath or difficulty breathing, fatigue, muscle or body aches, headache, new loss of taste or smell, sore throat, congestion or runny nose, nausea, vomiting, or diarrhea.
  • the method reduces or inhibits susceptibility to B. pertussis infection or pathology.
  • the composition is administered prior to, substantially contemporaneously with or following exposure to or infection of the subject with B. pertussis.
  • the composition is administered prior to, substantially contemporaneously with or following exposure to or infection of the subject with B. pertussis.
  • the composition is administered within 2-72 hours, 2-48 hours, 4-24 hours, 4-18 hours, or 6-12 hours after a symptom of B. pertussis infection or exposure develops.
  • the composition is administered prior to exposure to or infection of the subject with B. pertussis.
  • the present invention includes a peptide or peptides that are immunoprevalent or immunodominant in a bacteria obtained by a method consisting of, or consisting essentially of: obtaining an amino acid sequence of the bacteria; determining one or more sets of overlapping peptides spanning one or more bacteria antigen using unbiased selection; synthesizing one or more pools of bacteria peptides comprising the one or more sets of overlapping peptides; combining the one or more pools of bacteria peptides with Class I major histocompatibility proteins (MHC), Class II MHC, or both Class I and Class II MHC to form peptide-MHC complexes; contacting the peptide-MHC complexes with T cells from subjects exposed to the bacteria; determining which pools triggered cytokine release by the T cells; and deconvoluting from the pool of peptides that elicited cytokine release by the T cells, which peptide or peptides are immunoprevalent or immunodominant
  • MHC major histo
  • the bacteria is a Bordetella.
  • the Bordetella is B. pertussis.
  • the immunodominant peptides are selected from 1, 2 or more peptides selected from the amino acid sequences set forth in any one of Tables 1-20.
  • the immunodominant peptides are selected from 1, 2 or more peptides selected from the amino acid sequences set forth in those sequences set forth in any one of Tables 1-20.
  • the present invention includes a method of selecting an immunoprevalent or immunodominant peptide or protein of a bacteria comprising, consisting of, or consisting essentially of: obtaining an amino acid sequence of the bacteria; determining one or more sets of overlapping peptides spanning one or more bacteria antigen using unbiased selection; synthesizing one or more pools of bacteria peptides comprising the one or more sets of overlapping peptides; combining the one or more pools of bacteria peptides with Class I major histocompatibility proteins (MHC), Class II MHC, or both Class I and Class II MHC to form peptide-MHC complexes; contacting the peptide-MHC complexes with T cells from subjects exposed to the bacteria; determining which pools triggered cytokine release by the T cells; and deconvoluting from the pool of peptides that elicited cytokine release by the T cells, which peptide or peptides are immunoprevalent or immunodominant in the pool
  • MHC major histo
  • the bacteria is a Bordetella .
  • the Bordetella is B. pertussis.
  • the immunodominant peptides are selected from 1, 2 or more peptides selected from the amino acid sequences set forth in any one of Tables 1-20.
  • the immunodominant peptides are selected from 1, 2 or more peptides selected from the amino acid sequences set forth in those sequences set forth in any one of Tables 1-20.
  • the present invention includes a polynucleotide that expresses one or more peptides or proteins, comprising, consisting of, or consisting essentially of an amino acid sequence selected from any one of those sequences set forth in Tables 1-20, or a subsequence, portion, homologue, variant or derivative thereof; a fusion protein comprising one or more amino acid sequences selected from any one of those sequences set forth in Tables 1-20; or a pool of 2 or more or more peptides comprising, consisting of, or consisting essentially of amino acid sequences selected from any one of those sequences set forth in Tables 1-20.
  • the vector comprises the polynucleotide of claim that expresses one or more peptides or proteins, comprising, consisting of, or consisting essentially of an amino acid sequence selected from any one of those sequences set forth in Tables 1-20, or a subsequence, portion, homologue, variant or derivative thereof; a fusion protein comprising one or more amino acid sequences selected from any one of those sequences set forth in Tables 1-20; or a pool of 2 or more or more peptides comprising, consisting of, or consisting essentially of amino acid sequences selected from any one of those sequences set forth in Tables 1-20, a bacterial vector, or a host cell the comprises the same.
  • the present invention includes a polynucleotide that expresses one or more peptides or proteins comprising, consisting of, or consisting essentially of an amino acid sequence selected from any one of those sequences set forth in Tables 1-20, or a subsequence, portion, homologue, variant or derivative thereof; a fusion protein comprising one or more amino acid sequences selected from any one of those sequences set forth in Tables 1-20; or a pool of 2 or more peptides selected from any one of those sequences set forth in Tables 1-20.
  • the vector comprises the polynucleotide of claim that expresses one or more peptides or proteins comprising, consisting of, or consisting essentially of an amino acid sequence selected from any one of those sequences set forth in Tables 1-20, or a subsequence, portion, homologue, variant or derivative thereof; a fusion protein comprising one or more amino acid sequences selected from any one of those sequences set forth in Tables 1-20; or a pool of 2 or more peptides selected from any one of those sequences set forth in Tables 1-20, a bacterial vector, or a host cell that comprises the same.
  • FIGS. 1A-1D Schematics of BP whole genome-wide library screening. An example of the entire BP peptide library screening and epitope identification for a representative individual donor using AIM assay is shown.
  • FIG. 1A Screening of entire library organized in 133 pools of 188 15-mer peptides (MegaPools; MP).
  • FIG. IB Deconvolution of one positive representative MP (MP#39) into 8 pools of 22-24 individual peptides (MesoPools; MS).
  • FIGS. 2A-2D Large breadth of BP-specific CD4+ T cell responses in humans.
  • FIG. 2B Dominance of antigen response indicated by proportion of donors who responded to the specified number of ORFs.
  • FIG. 2C Breadth of epitope response ranked on the basis of % of total response (Black dashed line). Grey dotted lines indicate the top 50, 75 and 90 percent of total response and associated number of epitopes.
  • FIG. 2D Breadth of antigen response ranked on the basis of % of total response (Black dashed line). Grey dotted lines indicate the top 50, 75 and 90 percent of total response and associated number of ORFs.
  • FIG. 3 Immunodominance of BP specific-CD4+ T cell responses.
  • Overall map of CD4+ T cell responses by antigen (ORF) reactivity across the entire cohort (n 40) showing the position of each individual ORF identified across the aligned BP genome, using the Tohama I and D420 BP strains as reference.
  • Associated percentage of total response (all antigens recognized) for each ORF is indicated in y axis.
  • Each bar represents an individual ORF and annotation of specific antigens is shown (red - aP vaccine antigens; green - dominant non-aP vaccine antigens).
  • FIGS. 4A-4F aP and non-aP vaccine antigens are similarly recognized in aP- and wP-primed donors.
  • Graphs show comparison of responses between aP- and wP-primed donors in terms of (FIGS. 4A,4D) magnitude, (FIGS. 4B,4E) number of epitopes, and (FIGS. 4C,4F) number of ORFs for aP vaccine antigens (Grey triangles, upper panel) or non-aP vaccine antigens (Grey circles, bottom panel), respectively.
  • Bars represent geometric mean ⁇ geometric SD. p values calculated by Mann-Whitney statistical analysis are indicated.
  • FIGS. 5A-5L Sequence conservation is not a major driver of immunogenicity. Peptide homology amongst different BP strains or Bordetella genus was evaluated for the entire se of peptides tested in this study.
  • FIG. 5A percent of homology across different BP strains for non-reactive, subdominant or dominant non-aP vaccine derived peptides
  • FIG. 5B percent of dominant peptides across different BP strains for peptide conservation in non-Ap vaccine derived peptides.
  • FIG. 5C percent of variable peptides across different BP strains for non-reactive, subdominant or dominant non-Ap vaccine derived peptides.
  • FIG. 5D percent of homology across different BP strains for non-reactive, subdominant or dominant aP vaccine derived peptides.
  • FIG. 5E percent of dominant peptides across different BP strains for peptide conservation in aP vaccine derived peptides.
  • FIG. 5F percent of conserved peptides across different BP strains for non-reactive, subdominant or dominant aP vaccine derived peptides.
  • FIG. 5G percent of homology across different Bordetella for non-reactive, subdominant or dominant non-aP vaccine derived peptides.
  • FIG. 5H percent of dominant peptides across different Bordetella for peptide conservation in non-aP vaccine derived peptides.
  • FIG. 51 percent of variable peptides across different Bordetella for non- reactive, subdominant or dominant non-Ap vaccine derived peptides.
  • FIG. 5 J percent of homology across different Bordetella for non-reactive, subdominant or dominant aP vaccine derived peptides.
  • FIG. 5K percent of dominant peptides across different Bordetella for peptide conservation in aP vaccine derived peptides.
  • FIGS. 6A-6C Th2 polarization is specific to the aP vaccine antigens, in individuals originally primed with aP vaccine.
  • Antigen specific CD4+ T cell responses from aP and non-aP vaccine antigens were detected with PT(E)VAC and PT(E)R peptide pools respectively.
  • Black dotted lines represent the cut-off value associated with the threshold of positivity (TP) and percentage of donor recognition is indicated for each stimuli.
  • each circle represents a donor.
  • Tick black lines represent geometric mean ⁇ geometric SD, and p values were calculated by Mann- Whitney.
  • FIGS. 7A-7C non-aP vaccine antigen responses are not polarized as function of priming childhood vaccination.
  • Antigen specific CD4+ T cell responses from 15 individual non-aP vaccine antigens were detected with overlapping (O) peptide pools and represented as the sum of all MPs responses (PT(O)1-15) or as each individual MP response (ANT1-ANT15).
  • PT(E)VAC pool was used as control respectively.
  • FIG. 8 Schematic of BP genome-wide screening and summary of experimental design and strategy.
  • CD4+ T cell reactivity spanning the entire BP proteome was assayed with a library of 24,877 peptides in 3 sequential steps, directly ex vivo using a high throughput Activation Induced Marker (AIM) assay flow cytometry methodology.
  • AIM Activation Induced Marker
  • FIGS. 9A-9B Immunodominance is associated with both magnitude and donor recognition.
  • FIG. 9B Graph shows correlation between percentages of total magnitude and donor recognition. Each circle represents an individual ORF. R and p value expresses Spearman's rank correlation coefficient test.
  • CD 154+ CD4+ T cells Representative gating of reactive OX40+CD25+ and cytokine+(IFNy, IL-2, TNFa and IL-4) CD 154+ CD4+ T cells from donor PBMCs is shown. Briefly, for both AIM and ICS, mononuclear cells were gated out of all events followed by subsequent singlet gating. Live CD3+ cells were gated as Live/Dead-CD14-CD8-CD19-CD3+. Cells were then gated as CD4+CD3+.
  • antigen-specific cells were defined as OX40+CD25+ CD4+ T cells (AIM+) after antigen stimulation, and frequencies calculated as percent of total CD4+ T cells after background subtraction.
  • antigen-specific cytokine producing cells were defined as Cytokine+ and CD 154+ CD4+ T cells after antigen stimulation, and frequencies calculated as percent of total CD4+ T cells after background subtraction.
  • FIGS. 12A-12C Experimentally defined epitope pools (PT(E)VAC and PT(E)R) detect BP- specific responses in vaccinated subjects primed with acellular (aP) or whole-cell (wP) vaccines in childhood.
  • aP acellular
  • wP whole-cell
  • FIG. 13 Novel identified antigens have equal or higher reactivity than aP vaccine antigens.
  • FIG. 14 Overlapping peptide pools derived from the 19 most immunodominant antigens not contained in the aP vaccine, can be used to detect BP-specific responses.
  • Example 1 Bordetella pertussis (BP), the causative agent of whooping cough, infects human hosts' lungs and upper airways.
  • BP Bordetella pertussis
  • AP acellular
  • wP whole-cell
  • the antigens included are the filamentous hemagglutinin (FHA), pertactin (PRN), pertussis toxin (PT), and fimbrial proteins 2 and 3 (Fim2/3).
  • FHA filamentous hemagglutinin
  • PRN pertactin
  • PT pertussis toxin
  • Fim2/3 fimbrial proteins 2 and 3
  • the inventors have identified 19 antigens with reactivity as prominent or even more immunodominant than the aP vaccine antigens. These discoveries enabled the unique opportunity to generate epitope pools to characterize and discriminate responses specific to aP vaccine antigens from other immunodominant and immunogenic BP antigens not contained in aP vaccines. Specifically, and in addition to the [PT(E)VAC] epitope pool (Table 1), the inventors generated a new epitope pool of 170 experimentally defined peptides covering the most immunogenic peptides across the entire BP genome, and not containing aP vaccine antigens [PT(E)R] (Table 2).
  • the inventors further generated 19 sets of BP-specific CD4+ T cells epitopes pools using overlapping peptides covering the entire sequence of the novel antigens.
  • the inventors tested the sensitivity and performance of the BP- specific peptide pools after short-culture stimulation of PBMCs using activation induced marker (AIM) and intracellular cellular staining (ICS) assays.
  • PBMCs were obtained from 2 distinct cohorts including aP- or wP-primed individuals in childhood. The previous and new epitope pools will allow studying BP- specific responses in aP vaccination or boost settings and in the context of whole-cell vaccination schemes, exposition/colonization, infection, and human challenge studies.
  • Table 21 List of the immunodominant antigens with reactivity and frequency of recognition equal or better than aP vaccine antigens.
  • Example 2 Experimentally defined BP epitope pools can be used to detect BP-specific responses in vaccinated individuals.
  • BP-specific CD4+ T cell responses are detected in vaccinated individuals irrespective of the type of vaccine administered in childhood by activation induced marker (AIM) or intracellular cytokine staining (ICS) assays.
  • AIM activation induced marker
  • ICS intracellular cytokine staining
  • PT(E)VAC aP vaccine antigens
  • Table 1 previously experimentally defined (Bancroft et al., 2016) and thoroughly characterized (da Silva Antunes et al., 2018; da Silva Antunes et al., 2021 ; da Silva Antunes et al., 2020) was combined into a pool of epitopes (megapool) and used in short-culture stimulation of PBMCs in AIM or ICS assays.
  • the PT(E)VAC megapool includes peptides from 5 antigens [Filamentous hemagglutinin (FHA), pertactin (PRN), pertussis toxin (PT), and fimbrial proteins 2 and 3 (Fim2/3)], which are the only 5 antigens contained in the current acellular (aP) vaccine administered in the United States.
  • FHA hemagglutinin
  • PRN pertactin
  • PT pertussis toxin
  • Fimbrial proteins 2 and 3 Fimbrial proteins 2 and 3
  • This megapool was developed from the findings of an ongoing NIH Pertussis T cell contract (75N93019C00066) that spearheaded the identification of novel T cell epitope targets and novel immunogenic antigens (da Silva Antunes et al. in preparation). Interestingly, BP-specific reactivity was similar between the 2 megapools using an AIM assay after 24h of stimulation (FIG. 12A). Using a threshold for positivity of 100 AIM+ cells (indicated by the dotted line), 20/20 (100%) of donors showed positive responses after PT(E)VAC stimulation, and 16/20 (80%) of donors showed positive responses after (PT(E)R) stimulation.
  • Example 3 Focusing on individual selected antigens.
  • Individual antigen responses for the most immunodominant antigens (FIG. 13), identified in a BP genome-wide screening study (da Silva Antunes et al. in preparation) can also be detected in vaccinated individuals using the AIM assay (FIG. 14).
  • adenylate cyclase toxin (ACT), BvgS and BrkA, known to be strongly associated with virulence and BP infection in mouse models, were identified, as well as a type III secretion system protein (T3SS) or proteins involved in regulation, serum resistance, or DNA-binding.
  • T3SS type III secretion system protein
  • enzymes involved in roles such as cell wall and cell membrane assembly (D-alanyl-D-alanine carboxypeptidase and membrane protein insertase YidC, respectively), metabolic processes related with glycogen biosynthesis, oxidative phosphorylation or other catalytic processes were also highly reactive and elicited immunodominant responses.
  • the reactivity to these antigens is about 2-fold higher than for vaccine antigens (FIGS. 2A-2D).
  • antigen-specific responses for the 19 BP most immunodominant antigens not included in the aP vaccine can also be detected, altough with variable degree of reactivity and responsiveness.
  • the epitope pools for ACT (ANT10) and BrKA (ANT7) elected strong responses, and were the highest immunoreactive targets with 100% (20/20), and 80% (16/20) of positive donor response, respectively.
  • ACT ANT10
  • BrKA ANT7
  • PT(E)VAC PT(E)R
  • responses to all antigens were detected irrespectively of the vaccine administered in childhood immunization (not shown).
  • the inventors demonstrate that through the use of peptide pools described in this disclosure, detection and quantification of BP-specific T cells can be easily and rapidly accomplished with high sensitivity in vaccinated cohorts, irrespectively of the nature of their childhood BP vaccine immunization.
  • the newly developed BP human T cell epitope pools can be further used to measure T cell responses against BP colonization in naturally infected, and clinically diagnosed acute or convalescent cohorts.
  • the disclosure identifies novel immunogenic targets that are crucial in the design of vaccines for superior control of BP infection and induction of long-lasting protection.
  • Example 4 Experimental design for a genome-wide screen of Bordetella pertussis human T cell epitopes.
  • the library was screened for CD4+ T cell reactivity utilizing PBMCs collected in the 2013 to 2021 period from 40 participants, 21 males and 19 females, of 18 to 40 years of age. Based on clinical records and year of birth, 20 of the participants were immunized in childhood with a whole-cell Pertussis (wP) vaccine, and 20 were originally immunized in childhood with an acellular Pertussis (aP) vaccine.
  • WP whole-cell Pertussis
  • aP acellular Pertussis
  • Example 5 Large breadth of BP-specific CD4+ T cell responses in humans.
  • CD4+ T cell reactivity was assayed directly ex vivo using an Activation Induced Marker (AIM) assay (FIGS. 12A-12C), utilizing the combination of markers OX40+CD25+ (Dan et al., 2016), previously validated for epitope identification in the context of BP (da Silva Antunes et al., 2020).
  • AIM Activation Induced Marker
  • FIGS. 1A-1D An example of screening of the whole genome-wide library in a representative donor is shown in FIGS. 1A-1D. In this particular donor, 32 positive MPs were identified (FIG. 1A)
  • FIG. IB illustrates the deconvolution of one representative MP (MP#39), which yielded 3 positive MS.
  • FIG. 1A illustrates the deconvolution of one representative MP (MP#39), which yielded 3 positive MS.
  • FIG. ID shows the position of each individual epitope identified across the aligned BP genome, using the Tohama I and D420 BP strains as reference.
  • Example 6 Immunodominance in BP responses.
  • the overall magnitude of response and localization of each recognized ORF/antigen across the entire cohort was next visualized summing all the reactivity of individual epitopes across all donors on a linear map of the BP genome (FIG. 3).
  • the known aP vaccine antigens i.e., pertactin, PRN; two serotypes of fimbriae, Fim2/3; filamentous hemagglutinin, FHA; and pertussis toxin, PtTox
  • FHA was the antigen with the highest magnitude (2.12% of total response) and the most frequently recognized (45.0% of donors) of all the BP antigens.
  • PtTox (ORF 1-5) and PRN were also associated with a high reactivity (1.34% and 1.33% of total response, respectively) and high frequency of donor recognition (30.0% and 40%, respectively).
  • Fim 3 and Fim 2 had the lowest reactivity (0.24% and 0.09% of total response, respectively) and were the least recognized antigens among the aP vaccine antigens (12.5% and 5% of donors, respectively). Strikingly, the cumulative response of the 5 aP vaccine antigens only accounted for a minor fraction of the total CD4+ T cell response (5. 13% of the total response).
  • the 15 antigens eliciting same level of responses than the vaccine antigens included ORFs from enzymes involved in roles such as cell wall and cell membrane assembly (D-alanyl-D-alanine carboxypeptidase and membrane protein insertase YidC, respectively) and from a transporter protein (Fha C) that mediates the secretion of aP vaccine antigen FHA.
  • ORFs from enzymes involved in roles such as cell wall and cell membrane assembly (D-alanyl-D-alanine carboxypeptidase and membrane protein insertase YidC, respectively) and from a transporter protein (Fha C) that mediates the secretion of aP vaccine antigen FHA.
  • FHA transporter protein
  • ACT adenylate cyclase toxin
  • BvgS and BrkA which are associated with virulence and BP infection
  • Example 7 Similar recognition of aP and non-aP vaccine antigens as a function of priming vaccination in infancy.
  • Example 8 Sequence conservation and immunogenicity of BP peptides.
  • peptides were classified as variable ( ⁇ 75% of homology), intermediate (75-95% of homology), or conserved (>95% of homology). Finally, peptides were further segregated as derived from aP vaccine antigens, or from non-aP vaccine antigens.
  • Example 9 Phenotypes associated with recognition of non-aP vaccine antigens.
  • the inventors To characterize responses to the highest reactive non-aP vaccine antigens identified in this study, the inventors generated a pool encompassing 170 different epitopes (Dominant peptides tested positive in at least 2 donors with >0.06% total CD4+T cell response), and hereafter denominated PT(E)R (Table 2). As control, the inventors used a previously described MP (Bancroft et al., 2016; da Silva Antunes et al., 2018), containing epitopes exclusive from aP vaccine antigens [PT(E)VAC]. These MPs were tested in replicates of 3 independent experiments with PBMC from 20 subjects (10 aP and 10 wP).
  • CD4+ T cell responses to PT(E)VAC and PT(E)R were measured by intracellular cytokine staining (ICS) with phenotypic assessment of IFNy, TNFa, IL-2 and IL-4 expression among intracellular CD154+ (CD40L) cells in PBMC from an additional cohort of 40 subjects (20 aP and 20 wP).
  • ICS cytokine staining
  • CD154+ CD154+
  • Example 10 CD4+ T cell responses against individual non-aP vaccine antigens.
  • BP specific-CD4+ T cells in healthy young adults immunized with different pertussis vaccines are associated with a previously unrecognized and remarkable large breadth of responses.
  • Tens of ORFs and hundreds of different peptides were recognized in the 40 donors studied. This remarkable large breadth of T cell responses parallels the large breadth of responses observed for another bacterial species (Mycobacterium tuberculosis: (Lindestam Arlehamn et al., 2013)), and it is a testament to the fact that T cell responses have the capacity to recognize most if not all foreign proteins.
  • antigens included BrkA and ACT, known to play important roles in the pathogenesis of pertussis, as dominant targets of human T cell responses.
  • Other antigens were associated with regulation of gene expression (LysR family transcriptional regulator) or virulence (virulence sensor protein BvgS, YihY/virulence factor BrkB family protein or Filamentous hemagglutinin transporter protein FhaC, that mediates the secretion of aP vaccine antigen FHA).
  • virulence virulence sensor protein BvgS, YihY/virulence factor BrkB family protein or Filamentous hemagglutinin transporter protein FhaC, that mediates the secretion of aP vaccine antigen FHA.
  • PtTox translocate proteins and virulence factors
  • virulence factors and bacterial secretory systems appear to be frequently recognized by human T cell responses, possibly reflective of their role in infection (Lindestam Arlehamn et al., 2013; Rolan and Tsolis, 2008; Saikh et al., 2006; Shepherd and McLaren, 2020).
  • each individual tended to recognize a partially overlapping yet unique set of antigenic and epitope targets.
  • the reasons for this donor-donor heterogeneity are not apparent, but might include past infection history, differences in HLA types, and influences of the microbiome on the repertoire of recognition.
  • conservation amongst different Bordetella species is not a major driver of the dominantly recognized epitopes as in general, peptide homology or sequence conservation between Bordetella isolates was low, with only a small fraction ( ⁇ 3%) of the highly conserved peptides associated with dominant responses.
  • the majority of the Bordetella isolates studied are species non- infectious to humans, and therefore a high sequence homology devoid of relevance.
  • PBMC isolation PBMCs were isolated from whole blood by density gradient centrifugation according to manufacturer instructions (Ficoll-Hypaque, Amersham Biosciences, Uppsala, Sweden) and cryopreserved for further analysis.
  • Peptide prediction, synthesis, library assembly and pool preparation were derived from either BP whole-genome predictions from the Tohama I strain or from a set of 256 unique open reading frames (ORF) from the recent clinical isolate D420 strain and not contained in Tohama I strain. These 256 unique ORFs have been identified in the lab of Dr. Tod Merkel (unpublished findings or Ref?). BP genome-wide identification from Tohama I strain was performed by scanning for the presence of predicted HUA class II promiscuous binding peptides. MHC -peptide binding predictions were performed using publicly available tools hosted by the Immune Epitope Database (IEDB) Analysis Resource (Dhanda et al., 2019).
  • IEDB Immune Epitope Database
  • the prediction of peptides was established by the 7-allele HLA class II restricted method and by using peptides 15 residues in length and overlapping by 10 residues. Additional filtering using an epitope cluster analysis tool was performed to include unique peptides across all proteins with median percentile rank (cut-off of 10) predicted peptides for each antigen or at least 2 peptides per ORF. To remove redundant peptides, all peptides overlapping by 9 residues or more were placed into “variant clusters”. The most commonly occurring peptide was marked as the “representative” and the less-common peptides were marked as “variants”. Variants in each cluster were sorted by their alignment-start position and only synthesized once.
  • the inventors selected and synthesized a total of 24,876 peptides, spanning 3,305 unique ORFs.
  • the peptides were pooled and organized into a library of 1,064 MesoPools (MS) composed of 24 individual peptides, and also a library of 133 MegaPools (MP) composed of 8 MS. All individual peptides were synthesized by Mimotopes (Victoria, Australia) and resuspended to a final concentration of 1 mg/mL in DMSO.
  • MS MesoPools
  • MP MegaPools
  • FIGS. 9A-9B show the immunodominance is associated with both magnitude and donor recognition.
  • FIG. 9B Graph shows correlation between percentages of total magnitude and donor recognition. Each circle represents an individual ORF. R and p value expresses Spearman's rank correlation coefficient test.
  • Activation Induced Marker AIM
  • ICS Intracellular staining assays.
  • CD4+ T cell reactivity was assayed directly ex vivo using an Activation induced marker (AIM) assay utilizing the combination of markers OX40+CD25+ as previously described (Dan etal., 2016). This assay detects cells that are activated as a result of antigen-specific stimulation by staining antigen-experienced CD4+ T cells for TCR- dependent upregulation of 0X40 and CD25 (AIM25) after an optimal time of 18-24 h of culture.
  • AIM Activation Induced Marker
  • ICS Intracellular staining
  • cryopreserved PBMCs were thawed, and 1 x 10 6 cells/condition were immediately cultured together with peptide pools (2 pg/mL), individual peptides (10 pg/mL), or PHA (10 pg/mL; Roche, San Diego, CA) and DMSO as positive and negative controls, respectively, in 5% human serum (Gemini Bio-Products) for 24 h. All samples were acquired on a ZE5 cell analyzer (Biorad laboratories, Hercules, CA) and analyzed with FlowJo software (Tree Star, Ashland, OR). AIM+ CD4+ T cells data were calculated as percentage of cells per million of CD4+ T cells.
  • ICS intracellular cytokine staining
  • lymphocytes were gated, followed by single cells determination. T cells were gated for being positive to CD3 and negative for a Dump channel including in the same colors CD14, CD19 and Live/Dead staining. CD3+CD4+ were further gated based on a combination of each cytokine (IFNy, TNFa, IL-2, and IL-4) with CD40L (CD154). The total cytokine response and T cell functionality was calculated from Boolean gating of single cytokines that was applied to CD3+CD4+ cells.
  • the background was removed from the data by subtracting the average of the % of Cytokine+ cells plated in triplicate wells stimulated with DMSO.
  • CD4+ T cell cytokine responses were background subtracted individually and found positive only if fulfilling the criteria of an SI greater than 2 and above a threshold of positivity (TP) of 0.002% for overall CD4+Cytokine+ cells.
  • TP threshold of positivity
  • the TP for ICS was considered to be a positive response based on the median twofold standard deviation of T cell reactivity in negative DMSO controls.
  • peptides were further segregated as derived from aP vaccine antigens, or from non-aP vaccine antigens. Results were plotted as geomean of percent homology or relative percent of the number of peptides in each subset.
  • Statistical analysis Comparisons between groups were performed using the nonparametric two- tailed, and unpaired Mann-Whitney test or Kruskal-Wallis test adjusted with Dunn’s test for multiple comparisons. Spearman's rank correlation coefficient test was used for association analysis. Prism 8.0.1 (GraphPad, San Diego, CA, USA) was used for these calculations. Values pertaining to significance and correlation coefficient (R) are noted in the respective figure, and P ⁇ 0.05 defined as statistically significant.
  • Study approval This study was performed with approvals from the Institutional Review Board at La Jolla Institute for Immunology (protocols; VD-101-0513 and VD-059-0813). All participants provided written informed consent for participation and clinical medical history was collected and evaluated.
  • the term "gene” means the segment of DNA involved in producing a protein; it includes regions preceding and following the coding region (leader and trailer) as well as intervening sequences (introns) between individual coding segments (exons). The leader, the trailer as well as the introns include regulatory elements that are necessary during the transcription and the translation of a gene. Further, a “protein gene product” is a protein expressed from a particular gene. [0194] The word “expression” or “expressed” as used herein in reference to a gene means the transcriptional and/or translational product of that gene.
  • the level of expression of a DNA molecule in a cell may be determined on the basis of either the amount of corresponding mRNA that is present within the cell or the amount of protein encoded by that DNA produced by the cell.
  • the level of expression of non-coding nucleic acid molecules may be detected by standard PCR or Northern blot methods well known in the art. See, Sambrook et al., 1989 Molecular Cloning: A Laboratory Manual, 18.1- 18.88.
  • amino acid refers to naturally occurring and synthetic amino acids, as well as amino acid analogs and amino acid mimetics that function in a manner similar to the naturally occurring amino acids.
  • Naturally occurring amino acids are those encoded by the genetic code, as well as those amino acids that are later modified, e.g., hydroxyproline, y-carboxyglutamate, and O-phosphoserine.
  • Amino acid analogs refers to compounds that have the same basic chemical structure as a naturally occurring amino acid, i.e., an a carbon that is bound to a hydrogen, a carboxyl group, an amino group, and an R group, e.g., homoserine, norleucine, methionine sulfoxide, methionine methyl sulfonium. Such analogs have modified R groups (e.g. , norleucine) or modified peptide backbones, but retain the same basic chemical structure as a naturally occurring amino acid.
  • Amino acid mimetics refers to chemical compounds that have a structure that is different from the general chemical structure of an amino acid, but that functions in a manner similar to a naturally occurring amino acid.
  • the terms “non-naturally occurring amino acid” and “unnatural amino acid” refer to amino acid analogs, synthetic amino acids, and amino acid mimetics which are not found in nature.
  • Amino acids may be referred to herein by either their commonly known three letter symbols or by the one-letter symbols recommended by the IUPAC-IUB Biochemical Nomenclature Commission. Nucleotides, likewise, may be referred to by their commonly accepted single-letter codes.
  • polypeptide refers to a polymer of amino acid residues, wherein the polymer may, in embodiments, be conjugated to a moiety that does not consist of amino acids.
  • the terms apply to amino acid polymers in which one or more amino acid residue is an artificial chemical mimetic of a corresponding naturally occurring amino acid, as well as to naturally occurring amino acid polymers and non-naturally occurring amino acid polymers.
  • a “fusion protein” refers to a chimeric protein encoding two or more separate protein sequences that are recombinantly expressed as a single moiety.
  • Proteins and peptides include isolated and purified forms. Proteins and peptides also include those immobilized on a substrate, as well as amino acid sequences, subsequences, portions, homologues, variants, and derivatives immobilized on a substrate.
  • Proteins and peptides can be included in compositions, for example, a pharmaceutical composition.
  • a pharmaceutical composition is suitable for specific or non-specific immunotherapy, or is a vaccine composition.
  • Isolated nucleic acid including isolated nucleic acid
  • Cells expressing a protein or peptide are further provided. Such cells include eukaryotic and prokaryotic cells, such as mammalian, insect, fungal and bacterial cells.
  • Methods and uses and medicaments of proteins and peptides of the invention are included. Such methods, uses and medicaments include modulating immune activity of a cell against a pathogen, for example, a bacteria or bacteria.
  • peptide mimetic refers to protein-like chain designed to mimic a peptide or protein.
  • Peptide mimetics may be generated by modifying an existing peptide or by designing a compound that mimic peptides, including peptoids and [3-peptides.
  • Constantly modified variants applies to both amino acid and nucleic acid sequences. With respect to particular nucleic acid sequences, “conservatively modified variants” refers to those nucleic acids that encode identical or essentially identical amino acid sequences. Because of the degeneracy of the genetic code, a number of nucleic acid sequences will encode any given protein. For instance, the codons GCA, GCC, GCG and GCU all encode the amino acid alanine. Thus, at every position where an alanine is specified by a codon, the codon can be altered to any of the corresponding codons described without altering the encoded polypeptide. Such nucleic acid variations are "silent variations,” which are one species of conservatively modified variations.
  • Every nucleic acid sequence herein which encodes a polypeptide also describes every possible silent variation of the nucleic acid.
  • each codon in a nucleic acid except AUG, which is ordinarily the only codon for methionine, and TGG, which is ordinarily the only codon for tryptophan
  • TGG which is ordinarily the only codon for tryptophan
  • amino acid sequences one of skill will recognize that individual substitutions, deletions or additions to a nucleic acid, peptide, polypeptide, or protein sequence which alters, adds or deletes a single amino acid or a small percentage of amino acids in the encoded sequence is a "conservatively modified variant" where the alteration results in the substitution of an amino acid with a chemically similar amino acid. Conservative substitution tables providing functionally similar amino acids are well known in the art. Such conservatively modified variants are in addition to and do not exclude polymorphic variants, interspecies homologs, and alleles of the disclosure.
  • the following eight groups each contain amino acids that are conservative substitutions for one another: (1) Alanine (A), Glycine (G); (2) Aspartic acid (D), Glutamic acid (E); (3) Asparagine (N), Glutamine (Q); (4) Arginine (R), Lysine (K); (5) Isoleucine (I), Leucine (L), Methionine (M), Valine (V); (6) Phenylalanine (F), Tyrosine (Y), Tryptophan (W); (7) Serine (S), Threonine (T); and (8) Cysteine (C), Methionine (M) (see, e.g., Creighton, Proteins (1984)).
  • a "percentage of sequence identity” is determined by comparing two optimally aligned sequences over a comparison window, wherein the portion of the polynucleotide or polypeptide sequence in the comparison window may comprise additions or deletions (i.e. , gaps) as compared to the reference sequence (which does not comprise additions or deletions) for optimal alignment of the two sequences.
  • the percentage is calculated by determining the number of positions at which the identical nucleic acid base or amino acid residue occurs in both sequences to yield the number of matched positions, dividing the number of matched positions by the total number of positions in the window of comparison and multiplying the result by 100 to yield the percentage of sequence identity.
  • nucleic acids or polypeptide sequences refer to two or more sequences or subsequences that are the same or have a specified percentage of amino acid residues or nucleotides that are the same (i.e., about 60% identity, preferably 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or higher identity over a specified region, when compared and aligned for maximum correspondence over a comparison window or designated region) as measured using a BLAST or BLAST 2.0 sequence comparison algorithms with default parameters described below, or by manual alignment and visual inspection (see, e.g., NCBI web site ncbi.nlm.nih.gov/BLAST/ or the like).
  • sequences are then said to be “substantially identical.”
  • This definition also refers to, or may be applied to, the compliment of a test sequence.
  • the definition also includes sequences that have deletions and/or additions, as well as those that have substitutions.
  • the preferred algorithms can account for gaps and the like.
  • identity exists over a region that is at least about 25 amino acids or nucleotides in length, or more preferably over a region that is 50-100 amino acids or nucleotides in length.
  • amino acid or nucleotide base "position” is denoted by a number that sequentially identifies each amino acid (or nucleotide base) in the reference sequence based on its position relative to the N- terminus (or 5 '-end). Due to deletions, insertions, truncations, fusions, and the like that must be taken into account when determining an optimal alignment, in general the amino acid residue number in a test sequence determined by simply counting from the N-terminus will not necessarily be the same as the number of its corresponding position in the reference sequence.
  • a variant has a deletion relative to an aligned reference sequence
  • that insertion will not correspond to a numbered amino acid position in the reference sequence.
  • truncations or fusions there can be stretches of amino acids in either the reference or aligned sequence that do not correspond to any amino acid in the corresponding sequence.
  • multimer refers to a complex comprising multiple monomers (e.g., a protein complex) associated by noncovalent bonds.
  • the monomers be substantially identical monomers, or the monomers may be different.
  • the multimer is a dimer, a trimer, a tetramer, or a pentamer.
  • MHC Major Histocompatibility Complex
  • HLA human leucocyte antigens
  • MHC Class I or Class II multimers are well known in the art and include but are not limited to dimers, tetramers, pentamers, hexamers, heptamers and octamers.
  • MHC/peptide multimer refers to a stable multimeric complex composed of MHC protein(s) subunits loaded with a peptide of the present invention.
  • an MHC/peptide multimer include, but are not limited to, an MHC/peptide dimer, trimer, tetramer, pentamer or higher valency multimer.
  • MHC/peptide complex include, but are not limited to, an MHC/peptide dimer, trimer, tetramer, pentamer or higher valency multimer.
  • HLA human leukocyte antigens
  • Non-classical human MHC class I molecules such as HLA-E (homolog of mice Qa-lb) and MICA/B molecules are also encompassed by the present invention.
  • the MHC/peptide multimer is an HLA/peptide multimer selected from the group consisting of HLA-A/peptide multimer, HLA-B/peptide multimer, HLA-C/peptide multimer, HLA-E/peptide multimer, MICA/peptide multimer and MICB/peptide multimer.
  • HLA-DR HLA-DR
  • HLA-DP HLA-DP
  • HLA-DQ HLA-DQ
  • HLA-DQAl*01 HLA-DRBl*01
  • HLA-DRBl*03 HLA-DRBl*03
  • HLA-DRBl*03 non-classical human MHC class II molecules
  • HLA -DM and HL-DOA homolog in mice is H2-DM and H2-O
  • the MHC/peptide multimer is an HLA/peptide multimer selected from the group consisting of HLA-DP/peptide multimer, HLA-DQ/peptide multimer, HLA- DR/peptide multimer, HLA-DM/peptide multimer and HLA-DO/peptide multimer.
  • An MHC/peptide multimer may be a multimer where the heavy chain of the MHC is biotinylated, which allows combination as a tetramer with streptavidin. MHC -peptide tetramers have increased avidity for the appropriate T cell receptor (TCR) on T lymphocytes.
  • TCR T cell receptor
  • the multimers can also be attached to paramagnetic particles or magnetic beads to facilitate removal of non-specifically bound reporter and cell sorting. Multimer staining does not kill the labelled cells, thus, cell integrity is maintained for further analysis.
  • the MHC/peptide multimer of the present invention is particularly suitable for isolating and/or identifying a population of CD8+ T cells having specificity for the peptide of the present invention (in a flow cytometry assay).
  • the peptides or MHC class I or class II multimer as described herein is particularly suitable for detecting T cells specific for one or more peptides of the present invention.
  • the peptide(s) and/or the MHC/multimer complex of the present invention is particularly suitable for diagnosing Bordetella infection in a subject.
  • the method comprises obtaining a blood or PBMC sample obtained from the subject with an amount of a least peptide of the present invention and detecting at least one T cell displaying a specificity for the peptide.
  • Another diagnostic method of the present invention involves the use of a peptide of the present invention that is loaded on multimers as described above, so that the isolated CD8+ or CD4+ T cells from the subject are brought into contact with the multimers, at which the binding, activation and/or expansion of the T cells is measured.
  • the number of CD8+ and/or CD4+ cells binding specifically to the HLA-peptide multimer may be quantified by measuring the secretion of lymphokines/cytokines, division of the T cells, or standard flow cytometry methods, such as, for example, using fluorescence activated cell sorting (FACS).
  • FACS fluorescence activated cell sorting
  • the multimers can also be attached to paramagnetic ferrous or magnetic beads to facilitate removal of non-specifically bound reporter and cell sorting.
  • the MHC class I or class II peptide multimers as described herein can also be used as therapeutic agents.
  • the peptide and/or the MHC class I or class II peptide multimers of the present invention are suitable for treating or preventing a Bordetella infection in a subject.
  • the MHC Class I or Class II multimers can be administered in soluble form or loaded on nanoparticles.
  • antibody refers to a polypeptide encoded by an immunoglobulin gene or functional fragments thereof that specifically binds and recognizes an antigen.
  • the recognized immunoglobulin genes include the kappa, lambda, alpha, gamma, delta, epsilon, and mu constant region genes, as well as the myriad immunoglobulin variable region genes.
  • Light chains are classified as either kappa or lambda.
  • Heavy chains are classified as gamma, mu, alpha, delta, or epsilon, which in turn define the immunoglobulin classes, IgG, IgM, IgA, IgD and IgE, respectively.
  • the specified antibodies bind to a particular protein at least two times the background and more typically more than 10 to 100 times background.
  • Specific binding to an antibody under such conditions requires an antibody that is selected for its specificity for a particular protein.
  • polyclonal antibodies can be selected to obtain only a subset of antibodies that are specifically immunoreactive with the selected antigen and not with other proteins.
  • This selection may be achieved by subtracting out antibodies that cross-react with other molecules.
  • a variety of immunoassay formats may be used to select antibodies specifically immunoreactive with a particular protein.
  • solid-phase ELISA immunoassays are routinely used to select antibodies specifically immunoreactive with a protein (see, e.g., Harlow & Lane, Using Antibodies, A Laboratory Manual (1998) for a description of immunoassay formats and conditions that can be used to determine specific immunoreactivity).
  • Antibodies are large, complex molecules (molecular weight of -150,000 or about 1320 amino acids) with intricate internal structure.
  • a natural antibody molecule contains two identical pairs of polypeptide chains, each pair having one light chain and one heavy chain.
  • Each light chain and heavy chain in turn consists of two regions: a variable ("V") region involved in binding the target antigen, and a constant (“C") region that interacts with other components of the immune system.
  • the light and heavy chain variable regions come together in 3-dimensional space to form a variable region that binds the antigen (for example, a receptor on the surface of a cell).
  • Within each light or heavy chain variable region there are three short segments (averaging 10 amino acids in length) called the complementarity determining regions ("CDRs").
  • the six CDRs in an antibody variable domain fold up together in 3 -dimensional space to form the actual antibody binding site which docks onto the target antigen.
  • the position and length of the CDRs have been precisely defined by Kabat, E. et al., Sequences of Proteins of Immunological Interest, U.S. Department of Health and Human Services, 1983, 1987.
  • the part of a variable region not contained in the CDRs is called the framework ("FR"), which forms the environment for the CDRs.
  • antibody is used according to its commonly known meaning in the art. Antibodies exist, e.g., as intact immunoglobulins or as a number of well-characterized fragments produced by digestion with various peptidases. Thus, for example, pepsin digests an antibody below the disulfide linkages in the hinge region to produce F(ab)'2, a dimer of Fab which itself is a light chain joined to VH-CHI by a disulfide bond. The F(ab)'2 may be reduced under mild conditions to break the disulfide linkage in the hinge region, thereby converting the F(ab)'2 dimer into a Fab' monomer.
  • the Fab' monomer is essentially Fab with part of the hinge region (see Fundamental Immunology (Paul ed., 3d ed. 1993). While various antibody fragments are defined in terms of the digestion of an intact antibody, one of skill will appreciate that such fragments may be synthesized de novo either chemically or by using recombinant DNA methodology. Thus, the term antibody, as used herein, also includes antibody fragments either produced by the modification of whole antibodies, orthose synthesized de novo using recombinant DNA methodologies (e.g., single chain Fv) or those identified using phage display libraries (see, e.g., McCafferty et al., Nature 348:552-554 (1990)).
  • An exemplary immunoglobulin (antibody) structural unit comprises a tetramer.
  • Each tetramer is composed of two identical pairs of polypeptide chains, each pair having one “light” (about 25 kD) and one “heavy” chain (about 50-70 kD).
  • the N-terminus of each chain defines a variable region of about 100 to 110 or more amino acids primarily responsible for antigen recognition.
  • the terms variable light chain (VL) and variable heavy chain (VH) refer to these light and heavy chains respectively.
  • the Fc i.e., fragment crystallizable region
  • the Fc region is the “base” or "tail" of an immunoglobulin and is typically composed of two heavy chains that contribute two or three constant domains depending on the class of the antibody. By binding to specific proteins, the Fc region ensures that each antibody generates an appropriate immune response for a given antigen.
  • the Fc region also binds to various cell receptors, such as Fc receptors, and other immune molecules, such as complement proteins.
  • epitopes include but are not limited to a polypeptide and a nucleic acid encoding a polypeptide, wherein expression of the nucleic acid into a polypeptide is capable of stimulating an immune response when the polypeptide is processed and presented on a Major Histocompatibility Complex (MHC) molecule.
  • MHC Major Histocompatibility Complex
  • epitopes include peptides presented on the surface of cells non-covalently bound to the binding groove of Class I or Class II MHC, such that they can interact with T cell receptors and the respective T cell accessory molecules.
  • antigens and epitopes also apply when discussing the antigen binding portion of an antibody, wherein the antibody binds to a specific structure of the antigen.
  • Epitopes that are displayed by MHC on antigen presenting cells are cleavage peptides or products of larger peptide or protein antigen precursors.
  • protein antigens are often digested by proteasomes resident in the cell. Intracellular proteasomal digestion produces peptide fragments of about 3 to 23 amino acids in length that are then loaded onto the MHC protein. Additional proteolytic activities within the cell, or in the extracellular milieu, can trim and process these fragments further. Processing of MHC Class II epitopes generally occurs via intracellular proteases from the lysosomal/endosomal compartment.
  • the present invention includes, in one embodiment, pre- processed peptides that are attached to the anti-CD40 antibody (or fragment thereof) that directs the peptides against which an enhanced immune response is sought directly to antigen presenting cells.
  • the present invention includes methods for specifically identifying the epitopes within antigens most likely to lead to the immune response sought for the specific sources of antigen presenting cells and responder T cells.
  • T cell epitope refers to a specific amino acid that when present in the context of a Major or Minor Histocompatibility Complex provides a reactive site for a T cell receptor.
  • the T-cell epitopes or peptides that stimulate the cellular arm of a subject's immune system are short peptides of about 8-25 amino acids.
  • T-cell epitopes are recognized by T cells from animals that are immune to the antigen of interest.
  • These T-cell epitopes or peptides can be used in assays such as the stimulation of cytokine release or secretion or evaluated by constructing major histocompatibility (MHC) proteins containing or “presenting” the peptide.
  • MHC major histocompatibility
  • Such immunogenically active fragments are often identified based on their ability to stimulate lymphocyte proliferation in response to stimulation by various fragments from the antigen of interest.
  • the term “immunological response” refers to an antigen or composition is the development in a subject of a humoral and/or a cellular immune response to an antigen present in the composition of interest.
  • a “humoral immune response” refers to an immune response mediated by antibody molecules
  • a “cellular immune response” is one mediated by T-lymphocytes and/or other white blood cells.
  • CTL cytolytic T-cells
  • CTLs have specificity for peptide antigens that are presented in association with proteins encoded by the major histocompatibility complex (MHC) and expressed on the surfaces of cells.
  • MHC major histocompatibility complex
  • helper T-cells help induce and promote the destruction of intracellular microbes, or the lysis of cells infected with such microbes.
  • Another aspect of cellular immunity involves an antigenspecific response by helper T-cells.
  • Helper T-cells act to help stimulate the function, and focus the activity of, nonspecific effector cells against cells displaying peptide antigens in association with MHC molecules on their surface.
  • a “cellular immune response” also refers to the production of cytokines, chemokines and other such molecules produced by activated T-cells and/or other white blood cells, including those derived from CD4+ and CD8+ T-cells.
  • an immunological response may include one or more of the following effects: the production of antibodies by B-cells; and/or the activation of effector and/or suppressor T-cells and/or gamma-delta T-cells directed specifically to an antigen or antigens present in the composition or vaccine of interest.
  • These responses may serve to neutralize infectivity, and/or mediate antibody-complement, or antibody dependent cell cytotoxicity (ADCC) to provide protection to an immunized host.
  • ADCC antibody dependent cell cytotoxicity
  • Such responses can be determined using standard immunoassays and neutralization assays, well known in the art.
  • an “immunogenic composition” and “vaccine” refer to a composition that comprises an antigenic molecule where administration of the composition to a subject or patient results in the development in the subject of a humoral and/or a cellular immune response to the antigenic molecule of interest.
  • Vaccine refers to a composition that can provide active acquired immunity to and/or therapeutic effect (e.g., treatment) of a particular disease or a pathogen.
  • a vaccine typically contains one or more agents that can induce an immune response in a subject against a pathogen or disease, i.e., a target pathogen or disease.
  • the immunogenic agent stimulates the body’s immune system to recognize the agent as a threat or indication of the presence of the target pathogen or disease, thereby inducing immunological memory so that the immune system can more easily recognize and destroy any of the pathogen on subsequent exposure.
  • Vaccines can be prophylactic (e.g., preventing or ameliorating the effects of a future infection by any natural or pathogen) or therapeutic (e.g., reducing symptoms or aberrant conditions associated with infection).
  • the administration of vaccines is referred to vaccination.
  • a vaccine composition can provide nucleic acid, e.g., mRNA that encodes antigenic molecules (e.g., peptides) to a subject.
  • the nucleic acid that is delivered via the vaccine composition in the subject can be expressed into antigenic molecules and allow the subject to acquire immunity against the antigenic molecules.
  • the vaccine composition can provide mRNA encoding antigenic molecules that are associated with a certain pathogen, e.g., one or more peptides that are known to be expressed in the pathogen (e.g., pathogenic bacterium or bacteria).
  • nucleic acid molecules specifically polynucleotides, primary constructs and/or mRNA that encode one or more polynucleotides that express one or more peptides or proteins, comprising, consisting of, or consisting essentially of an amino acid sequence selected from any one of those sequences set forth in Tables 1-20, or a subsequence, portion, homologue, variant or derivative thereof for use in immune modulation.
  • nucleic acid refers to any compound and/or substance that comprise a polymer of nucleotides, referred to herein as polynucleotides.
  • nucleic acids or polynucleotides of the invention include, but are not limited to, ribonucleic acids (RNAs), deoxyribonucleic acids (DNAs), threose nucleic acids (TNAs), glycol nucleic acids (GNAs), peptide nucleic acids (PNAs), locked nucleic acids (LNAs), including diastereomers of LNAs, functionalized LNAs, or hybrids thereof.
  • RNAs ribonucleic acids
  • DNAs deoxyribonucleic acids
  • TAAs threose nucleic acids
  • GNAs glycol nucleic acids
  • PNAs peptide nucleic acids
  • LNAs locked nucleic acids
  • One method of immune modulation of the present invention includes direct or indirect gene transfer, i.e., local application of a preparation containing the one or more polynucleotides (DNA, RNA, mRNA, etc.) that expresses the one or more peptides or proteins, comprising, consisting of, or consisting essentially of an amino acid sequence selected from any one of those sequences set forth in Tables 1-20, or a subsequence, portion, homologue, variant or derivative thereof.
  • a variety of well-known vectors can be used to deliver to cells the one or more polynucleotides or the peptides or proteins expressed by the polynucleotides, including but not limited to adenobacterial vectors and adeno-associated vectors.
  • naked DNA, liposome delivery methods, or other novel vectors developed to deliver the polynucleotides to cells can also be beneficial.
  • promoters can be used to drive peptide or protein expression, including but not limited to endogenous promoters, constitutive promoters (e.g., cytomegalobacteria, adenobacteria, or SV40), inducible promoters (e.g., a cytokine promoter such as the interleukin- 1, tumor necrosis factor-alpha, or interleukin-6 promoter), and tissue specific promoters to express the immunogenic peptides or proteins of the present invention.
  • constitutive promoters e.g., cytomegalobacteria, adenobacteria, or SV40
  • inducible promoters e.g., a cytokine promoter such as the interleukin- 1, tumor necrosis factor-alpha, or interleukin-6 promoter
  • tissue specific promoters e.g., a cytokine promoter such as the interleukin- 1, tumor necrosis factor-alpha, or interleuk
  • the immunization may include adenobacteria, adeno-associated bacteria, herpes bacteria, vaccinia bacteria, retrobacteriaes, or other bacterial vectors with the appropriate tropism for cells likely to present the antigenic peptide(s) or protein(s) may be used as a gene transfer delivery system for a therapeutic peptide(s) or protein(s), comprising, consisting of, or consisting essentially of an amino acid sequence selected from any one of those sequences set forth in Tables 1-20, or a subsequence, portion, homologue, variant or derivative thereof, gene expression construct.
  • Bacterial vectors which do not require that the target cell be actively dividing are particularly useful when the cells are accumulating, but not proliferative.
  • Numerous vectors useful for this purpose are generally known (Miller, Human Gene Therapy 15-14, 1990; Friedman, Science 244: 1275-1281, 1989; Eglitis and Anderson, BioTechniques 6:608-614, 1988; Tolstoshev and Anderson, Current Opinion in Biotechnology 1:55-61, 1990; Sharp, The Lancet 337: 1277-1278, 1991; Cometta et al., Nucleic Acid Research and Molecular Biology 36:311-322, 1987; Anderson, Science 226:401-409, 1984; Moen, Blood Cells 17:407-416, 1991; and Miller and Rosman, Bio Techniques 7:980-990, 1989; Le Gal La Salle et al., Science 259:988-990, 1993; and Johnson, Chest 107:77S-83S, 1995).
  • the immunization may also include inserting the one or more polynucleotides (DNA, RNA, mRNA, etc.) that express the one or more peptides or proteins, comprising, consisting of, or consisting essentially of an amino acid sequence selected from any one of those sequences set forth in Tables 1-20, or a subsequence, portion, homologue, variant or derivative thereof into the bacterial vector, along with another gene which encodes the ligand for a receptor on a specific target cell, for example, such that the vector is now target specific.
  • Bacterial vectors can be made target specific by attaching, for example, a sugar, a glycolipid, or a protein. Targeting can also be accomplished by using an antibody to target the bacterial vector.
  • Bacterial or non-bacterial approaches may also be employed for the introduction of one or more therapeutic polynucleotides that express the one or more peptides or proteins, comprising, consisting of, or consisting essentially of an amino acid sequence selected from any one of those sequences set forth in Tables 1-20, or a subsequence, portion, homologue, variant or derivative thereof, into polynucleotide- encoding polynucleotide into antigen presenting cells.
  • the polynucleotides may be DNA, RNA, mRNA that directly encode the one or more peptides or proteins of the present invention, or may be introduced as part of an expression vector.
  • an immunization includes colloidal dispersion systems that include macromolecule complexes, nanocapsules, microspheres, beads, and lipid-based systems including oil-in- water emulsions, micelles, mixed micelles, and liposomes and the one or more polynucleotides that express the one or more peptides or proteins, comprising, consisting of, or consisting essentially of an amino acid sequence selected from any one of those sequences set forth in Tables 1-20, or a subsequence, portion, homologue, variant or derivative thereof.
  • a colloidal system for use with the present invention is a liposome.
  • Liposomes are artificial membrane vesicles which are useful as delivery vehicles in vitro and in vivo. It has been shown that large unilamellar vesicles (LUV), which range in size from 0.2-4.0 micrometers that can encapsulate a substantial percentage of an aqueous buffer containing large macromolecules. RNA, DNA and intact virions can be encapsulated within the aqueous interior and be delivered to cells in a biologically active form (Fraley, et al., Trends Biochem. Sci., 6:77, 1981). In addition to mammalian cells, liposomes have been used for delivery of polynucleotides in plant, yeast and bacterial cells.
  • LUV large unilamellar vesicles
  • a liposome In order for a liposome to be an efficient gene transfer vehicle, the following characteristics should be present: (Zakut and Givol, supra) encapsulation of the genes of interest at high efficiency while not compromising their biological activity; (Feamhead, et al., supra) preferential and substantial binding to a target cell in comparison to non-target cells; (Korsmeyer, S. J., supra) delivery of the aqueous contents of the vesicle to the target cell cytoplasm at high efficiency; and (Kinoshita, et al., supra) accurate and effective expression of genetic information (Mannino, et al., Bio Techniques, 6:682, 1988).
  • the composition for immunizing the subject or patient may, in certain embodiments comprise a combination of phospholipid, particularly high-phase-transition-temperature phospholipids, usually in combination with steroids, especially cholesterol. Other phospholipids or other lipids may also be used.
  • the physical characteristics of liposomes depend on pH, ionic strength, and the presence of divalent cations.
  • the targeting of liposomes can be classified based on anatomical and mechanistic factors. Anatomical classification is based on the level of selectivity, for example, organ-specific, cell-specific, and organelle-specific. Mechanistic targeting can be distinguished based upon whether it is passive or active.
  • Passive targeting utilizes the natural tendency of liposomes to distribute to cells of the reticuloendothelial system (RES) in organs which contain sinusoidal capillaries.
  • Active targeting involves alteration of the liposome by coupling the liposome to a specific ligand such as a monoclonal antibody, sugar, glycolipid, or protein, or by changing the composition or size of the liposome in order to achieve targeting to organs and cell types other than the naturally occurring sites of localization, specifically, cells that can become infected with a Bordetella or interact with the proteins, peptides, and/or gene products of Bordetella, e.g., immune cells.
  • a specific ligand such as a monoclonal antibody, sugar, glycolipid, or protein
  • the immune modulating polynucleotide construct, composition, or formulation is preferably applied to a site that will enhance the immune response.
  • the immunization may be intramuscular, intraperitoneal, enteral, parenteral, intranasal, intrapulmonary, or subcutaneous.
  • polynucleotide expression is directed from any suitable promoter (e.g., the human cytomegalobacteria, simian bacteria 40, actin or adenobacteria constitutive promoters; or the cytokine or metalloprotease promoters for activated synoviocyte specific expression).
  • the immune modifying peptide(s) or protein(s) include polynucleotides, constructs and/or mRNAs that express the one or more polynucleotides that express the one or more peptides or proteins, comprising, consisting of, or consisting essentially of an amino acid sequence selected from any one of those sequences set forth in Tables 1-20, or a subsequence, portion, homologue, variant or derivative thereof, that are designed to improve one or more of the stability and/or clearance in tissues, uptake and/or kinetics, cellular access by the peptide(s) or protein(s), translational, mRNA half-life, translation efficiency, immune evasion, protein production capacity, accessibility to circulation, peptide(s) or protein(s) half-life and/or presentation in the context of MHC on antigen presenting cells.
  • the present invention contemplates immunization for use in both active and passive immunization embodiments.
  • Immunogenic compositions proposed to be suitable for use as a vaccine, may be prepared most readily directly from immunogenic peptides, proteins, monomers, multimers and/or peptide-MHC complexes prepared in a manner disclosed herein.
  • the antigenic material is generally processed to remove undesired contaminants, such as, small molecular weight molecules, incomplete proteins, or when manufactured in plant cells, plant components such as cell walls, plant proteins, and the like. Often, these immunizations are lyophilized for ease of transport and/or to increase shelf-life and can then be more readily dissolved in a desired vehicle, such as saline.
  • immunizations also referred to as vaccines
  • the preparation of immunizations that contain the immunogenic proteins of the present invention as active ingredients is generally well understood in the art, as exemplified by United States Letters Patents 4,608,251; 4,601,903; 4,599,231; 4,599,230; 4,596,792; and 4.578,770, all incorporated herein by reference.
  • immunizations are prepared as injectables.
  • the immunizations can be a liquid solution or suspension but may also be provided in a solid form suitable for solution in, or suspension in, liquid prior to injection may also be prepared.
  • the preparation may also be emulsified.
  • the active immunogenic ingredient is often mixed with excipients that are pharmaceutically acceptable and compatible with the active ingredient.
  • excipients are, for example, water, saline, dextrose, glycerol, ethanol, buffers, or the like and combinations thereof.
  • the immunization may contain minor amounts of auxiliary substances such as wetting or emulsifying agents, pH buffering agents, or adjuvants which enhance the effectiveness of the vaccines.
  • the immunization is/are administered in a manner compatible with the dosage formulation, and in such amount as will be therapeutically effective and immunogenic.
  • the quantity to be administered depends on the subject to be treated, including, e.g., the capacity of the individual's immune system to synthesize antibodies, and the degree of protection desired.
  • Precise amounts of active ingredient required to be administered depend on the judgment of the practitioner. However, suitable dosage ranges are of the order of several hundred micrograms active ingredient per vaccination. Suitable regimes for initial administration and booster shots are also variable but are typified by an initial administration followed by subsequent inoculations or other administrations.
  • the manner of application of the immunization may be varied widely. Any of the conventional methods for administration of a vaccine are applicable. These are believed to also include oral application on a solid physiologically acceptable base or in a physiologically acceptable dispersion, parenterally, by injection or the like. The dosage of the vaccine will depend on the route of administration and will vary according to the size of the host.
  • Various methods of achieving adjuvant effect for the vaccine includes use of agents such as aluminum hydroxide or phosphate (alum), commonly used as 0.05 to 0.1 percent solution in phosphate buffered saline, admixture with synthetic polymers of sugars (Carbopol) used as 0.25 percent solution, aggregation of the protein in the vaccine by heat treatment with temperatures ranging between 70° to 101°C for 30 second to 2-minute periods respectively. Aggregation by reactivating with pepsin treated (Fab) antibodies to albumin, mixture with bacterial cells such as C.
  • agents such as aluminum hydroxide or phosphate (alum), commonly used as 0.05 to 0.1 percent solution in phosphate buffered saline, admixture with synthetic polymers of sugars (Carbopol) used as 0.25 percent solution, aggregation of the protein in the vaccine by heat treatment with temperatures ranging between 70° to 101°C for 30 second to 2-minute periods respectively. Aggregation by reactivating with pepsin treated (Fab)
  • parvum or endotoxins or lipopolysaccharide components of gram-negative bacteria emulsion in physiologically acceptable oil vehicles such as mannide mono-oleate (Aracel A) or emulsion with 20 percent solution of a perfluorocarbon (Fluosol-DA) used as a block substitute may also be employed.
  • physiologically acceptable oil vehicles such as mannide mono-oleate (Aracel A) or emulsion with 20 percent solution of a perfluorocarbon (Fluosol-DA) used as a block substitute may also be employed.
  • the vaccine will be desirable to have multiple administrations of the vaccine, usually not exceeding six to ten immunizations, more usually not exceeding four immunizations and preferably one or more, usually at least about three immunizations.
  • the immunizations will normally be at from two to twelve-week intervals, more usually from three to five-week intervals. Periodic boosters at intervals of 1- 5 years, usually three years, will be desirable to maintain protective levels of the antibodies.
  • the course of the immunization may be followed by assays for antibodies for the supernatant antigens.
  • the assays may be performed by labeling with conventional labels, such as radionuclides, enzymes, fluorescent agents, and the like.
  • Plant cloning vectors Clontech Laboratories, Inc., Palo-Alto, Calif., and Pharmacia LKB Biotechnology, Inc., Pistcataway, N.J.; Hood, E., et al., J. Bacteriol. 168: 1291-1301 (1986); Nagel, R., et al., FEMS Microbiol. Lett. 67:325 (1990); An, et al., “Binary Vectors”, and others in Plant Molecular Biology Manual A3: 1-20 (1988); Miki, B. L. A., et al., pp.
  • the term “effective amount” or “effective dose” refers to that amount of the peptide or protein T cell epitopes of the invention sufficient to induce immunity, to prevent and/or ameliorate an infection or to reduce at least one symptom of an infection and/or to enhance the efficacy of another dose of peptide or protein T cell epitopes.
  • An effective dose may refer to the amount of peptide or protein T cell epitopes sufficient to delay or minimize the onset of an infection.
  • An effective dose may also refer to the amount of peptide or protein T cell epitopes that provides a therapeutic benefit in the treatment or management of an infection.
  • an effective dose is the amount with respect to peptide or protein T cell epitopes of the invention alone, or in combination with other therapies, that provides a therapeutic benefit in the treatment or management of an infection.
  • An effective dose may also be the amount sufficient to enhance a subject's (e.g., a human's) own immune response against a subsequent exposure to an infectious agent.
  • Levels of immunity can be monitored, e.g., by measuring amounts of neutralizing secretory and/or serum antibodies, e.g., by plaque neutralization, complement fixation, enzyme-linked immunosorbent, or microneutralization assay.
  • an “effective dose” is one that prevents disease and/or reduces the severity of symptoms.
  • a “reduction” of a symptom or symptoms means decreasing of the severity or frequency of the symptom(s), or elimination of the symptom(s).
  • a “prophylactically effective amount” of a drug is an amount of a drug that, when administered to a subject, will have the intended prophylactic effect, e.g., preventing or delaying the onset (or reoccurrence) of an injury, disease, pathology or condition, or reducing the likelihood of the onset (or reoccurrence) of an injury, disease, pathology, or condition, or their symptoms, in this case, an infectious disease, and more particularly, a Bordetella infection.
  • a prophylactically effective amount may be administered in one or more administrations.
  • Guidance can be found in the literature for appropriate dosages for given classes of pharmaceutical products. For example, for the given parameter, an effective amount will show an increase or decrease of at least 5%, 10%, 15%, 20%, 25%, 40%, 50%, 60%, 75%, 80%, 90%, or at least 100%. Efficacy can also be expressed as “-fold” increase or decrease. For example, a therapeutically effective amount can have at least a 1.2-fold, 1.5-fold, 2-fold, 5-fold, or more effect over a control.
  • the term “immune stimulator” refers to a compound that enhances an immune response via the body's own chemical messengers (cytokines). These molecules comprise various cytokines, lymphokines and chemokines with immunostimulatory, immunopotentiating, and pro- inflammatory activities, such as interferons, interleukins (e.g., IL-1, IL-2, IL-3, IL-4, IL-12, IL-13); growth factors (e.g., granulocyte -macrophage (GM)-colony stimulating factor (CSF)); and other immunostimulatory molecules, such as macrophage inflammatory factor, Flt3 ligand, B7.1; B7.2, etc.
  • the immune stimulator molecules can be administered in the same formulation as peptide or protein T cell epitopes s of the invention, or can be administered separately. Either the protein or an expression vector encoding the protein can be administered to produce an immunostimulatory effect.
  • the term “protective immune response” or “protective response” refers to an immune response mediated by antibodies against an infectious agent, which is exhibited by a vertebrate (e.g., a human), which prevents or ameliorates an infection or reduces at least one symptom thereof.
  • a vertebrate e.g., a human
  • Peptide and protein T cell epitopes of the invention can stimulate the production of antibodies that, for example, neutralize infectious agents, blocks infectious agents from entering cells, blocks replication of said infectious agents, and/or protect host cells from infection and destruction.
  • the term can also refer to an immune response that is mediated by T-lymphocytes and/or other white blood cells against an infectious agent, exhibited by a vertebrate (e.g., a human), that prevents or ameliorates flavibacteria infection or reduces at least one symptom thereof.
  • a vertebrate e.g., a human
  • Peptide and protein T cell epitopes of the invention can stimulate the T cell responses that, for example, neutralize infectious agents, kill bacteria infected cells, blocks infectious agents from entering cells, blocks replication of said infectious agents, and/or protect host cells from infection and destruction.
  • biological sample refers to materials obtained from or derived from a subject or patient.
  • a biological sample includes sections of tissues such as biopsy and autopsy samples, and frozen sections taken for histological purposes.
  • samples include bodily fluids such as blood and blood fractions or products (e.g., serum, plasma, platelets, red blood cells, and the like), sputum, tissue, cultured cells (e.g., primary cultures, explants, and transformed cells) stool, urine, synovial fluid, joint tissue, synovial tissue, synoviocytes, fibroblast-like synoviocytes, macrophage -like synoviocytes, immune cells, hematopoietic cells, fibroblasts, macrophages, T cells, etc.
  • bodily fluids such as blood and blood fractions or products (e.g., serum, plasma, platelets, red blood cells, and the like), sputum, tissue, cultured cells (e.g., primary cultures, explants, and transformed cells) stool, urine, synovial fluid, joint tissue
  • a biological sample is typically obtained from a eukaryotic organism, such as a mammal such as a primate e.g., chimpanzee or human; cow; dog; cat; a rodent, e.g., guinea pig, rat, mouse; rabbit; or a bird; reptile; or fish.
  • a mammal such as a primate e.g., chimpanzee or human; cow; dog; cat; a rodent, e.g., guinea pig, rat, mouse; rabbit; or a bird; reptile; or fish.
  • a “cell” refers to a cell carrying out metabolic or other function sufficient to preserve or replicate its genomic DNA.
  • a cell can be identified by well-known methods in the art including, for example, presence of an intact membrane, staining by a particular dye, ability to produce progeny or, in the case of a gamete, ability to combine with a second gamete to produce a viable offspring.
  • Cells may include prokaryotic and eukaryotic cells.
  • Prokaryotic cells include but are not limited to bacteria.
  • Eukaryotic cells include but are not limited to yeast cells and cells derived from plants and animals, for example mammalian, insect (e.g., spodoptera) and human cells. Cells may be useful when they are naturally nonadherent or have been treated not to adhere to surfaces, for example by trypsinization.
  • the term "contacting” is used in accordance with its plain ordinary meaning and refers to the process of allowing at least two distinct species to become sufficiently proximal to react, interact or physically touch. It should be appreciated, however, the resulting reaction product can be produced directly from a reaction between the added reagents or from an intermediate from one or more of the added reagents which can be produced in the reaction mixture.
  • the term “contacting” may include allowing two species to react, interact, or physically touch, wherein the two species may be, for example, an amino acid sequence, protein, or peptide as provided herein and an immune cell, such as a T cell.
  • a "control" sample or value refers to a sample that serves as a reference, usually a known reference, for comparison to a test sample.
  • a test sample can be taken from a test condition, e.g., in the presence of a test compound, and compared to samples from known conditions, e.g., in the absence of the test compound (negative control), or in the presence of a known compound (positive control).
  • a control can also represent an average value gathered from a number of tests or results.
  • controls can be designed for assessment of any number of parameters.
  • a control can be devised to compare therapeutic benefit based on pharmacological data (e.g., half-life) or therapeutic measures (e.g., comparison of side effects).
  • pharmacological data e.g., half-life
  • therapeutic measures e.g., comparison of side effects
  • One of skill in the art will understand which controls are valuable in a given situation and be able to analyze data based on comparisons to control values. Controls are also valuable for determining the significance of data. For example, if values for a given parameter are widely variant in controls, variation in test samples will not be considered as significant.
  • modulator refers to a composition that increases or decreases the level of a target molecule or the function of a target molecule or the physical state of the target of the molecule relative to the absence of the modulator.
  • modulate is used in accordance with its plain ordinary meaning and refers to the act of changing or varying one or more properties. “Modulation” refers to the process of changing or varying one or more properties. For example, as applied to the effects of a modulator on a target protein, to modulate means to change by increasing or decreasing a property or function of the target molecule or the amount of the target molecule.
  • a disease e.g. a protein associated disease, a cancer (e.g., cancer, inflammatory disease, autoimmune disease, or infectious disease)
  • a disease e.g. cancer, inflammatory disease, autoimmune disease, or infectious disease
  • the disease e.g. cancer, inflammatory disease, autoimmune disease, or infectious disease
  • a symptom of the disease is caused by (in whole or in part) the substance or substance activity or function.
  • a causative agent could be a target for treatment of the disease.
  • aberrant refers to different from normal. When used to describe enzymatic activity or protein function, aberrant refers to activity or function that is greater or less than a normal control or the average of normal non-diseased control samples. Aberrant activity may refer to an amount of activity that results in a disease, wherein returning the aberrant activity to a normal or nondisease-associated amount (e.g., by administering a compound or using a method as described herein), results in reduction of the disease or one or more disease symptoms.
  • subject refers to a living organism who is at risk of or prone to having a disease or condition, or who is suffering from a disease or condition that can be treated by administration of a composition or pharmaceutical composition as provided herein.
  • Non-limiting examples include humans and other primates, but also includes non-human primates such as chimpanzees and other apes and monkey species; farm animals such as cattle, sheep, pigs, goats and horses; domestic mammals such as dogs and cats; laboratory animals including rodents such as mice, rats and guinea pigs; birds, including domestic, wild and game birds such as chickens, turkeys and other gallinaceous birds, ducks, geese, and the like.
  • the term does not denote a particular age. Thus, both adult and newborn individuals are intended to be covered.
  • the system described above is intended for use in any of the above vertebrate species, since the immune systems of all of these vertebrates operate similarly.
  • the terms “disease” or “condition” refer to a state of being or health status of a patient or subject capable of being treated with a compound, pharmaceutical composition, or method provided herein.
  • a patient or subject is human.
  • the disease is Bordetella infection.
  • the disease is B. pertussis infection.
  • the disease is whooping cough.
  • “treatment” or “treating,” or “palliating” or “ameliorating” are used interchangeably herein. These terms refer to an approach for obtaining beneficial or desired results including but not limited to therapeutic benefit and/or a prophylactic benefit.
  • compositions may be administered to a patient at risk of bacterial infection, of developing a particular disease, or to a patient reporting one or more of the physiological symptoms of a disease, even though a diagnosis of this disease may not have been made.
  • Treatment includes preventing the infection or disease, that is, causing the clinical symptoms of the disease not to develop by administration of a protective composition prior to infection or the induction of the disease; suppressing the disease, that is, causing the clinical symptoms of the disease or infection not to develop by administration of a protective composition after the inductive event or infection but prior to the clinical appearance or reappearance of the disease; inhibiting the disease, that is, arresting the development of clinical symptoms by administration of a protective composition after their initial appearance; preventing re-occurring of the disease and/or relieving the disease, that is, causing the regression of clinical symptoms by administration of a protective composition after their initial appearance.
  • Treatment can also refer to any of (i) the prevention of infection or reinfection, as in a traditional vaccine, (ii) the reduction or elimination of symptoms, and (iii) the substantial or complete elimination of the pathogen in question. Treatment may be affected prophylactically (prior to infection) or therapeutically (following infection).
  • treatment refers to a method of reducing the effects of one or more symptoms of infection with a Bordetella.
  • treatment can refer to a 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100% reduction in the severity of an established infection, disease, condition, or symptom of the infection, disease or condition.
  • a method for treating a disease is considered to be a treatment if there is a 10% reduction in one or more symptoms of the disease in a subject as compared to a control.
  • the reduction can be a 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 100%, or any percent reduction in between 10% and 100% as compared to native or control levels. It is understood that treatment does not necessarily refer to a cure or complete ablation of the disease, condition, or symptoms of the disease or condition and/or complete prevention of infection. Further, as used herein, references to decreasing, reducing, or inhibiting include a change of 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90% or greater as compared to a control level and such terms can include but do not necessarily include complete elimination.
  • diagnosis refers to recognition of an infection, disease or condition by signs and symptoms. Diagnosing can refer to determination of whether a subject has an infection or disease. Diagnosis may refer to determination of the type of disease or condition a subject has or the type of bacteria the subject is infected with.
  • Diagnostic agents provided herein include any such agent, which are well-known in the relevant art.
  • imaging agents include fluorescent and luminescent substances, including, but not limited to, a variety of organic or inorganic small molecules commonly referred to as "dyes,” “labels,” or “indicators.” Examples include fluorescein, rhodamine, acridine dyes, Alexa dyes, and cyanine dyes.
  • Enzymes that may be used as imaging agents in accordance with the embodiments of the disclosure include, but are not limited to, horseradish peroxidase, alkaline phosphatase, acid phosphatase, glucose oxidase, [3-galactosidase, [3- glucoronidase or [3-lactamase. Such enzymes may be used in combination with a chromogen, a Anorogenic compound or a luminogenic compound to generate a detectable signal.
  • the peptide(s) or protein(s) of the present invention can also be used in binding assays including, but are not limited to, immunoassays such as competitive and non-competitive assay systems using techniques such as western blots, radioimmunoassays, ELISA (enzyme linked immunosorbent assay), "sandwich” immunoassays, Meso Scale Discovery (MSD, Gaithersburg, Md.), immunoprecipitation assays, ELISPOT, precipitin reactions, gel diffusion precipitin reactions, immunodiffusion assays, agglutination assays, complement-fixation assays, immunoradiometric assays, fluorescent immunoassays, and protein A immunoassays.
  • immunoassays such as competitive and non-competitive assay systems using techniques such as western blots, radioimmunoassays, ELISA (enzyme linked immunosorbent assay), "sandwich” immunoassays, Meso
  • Radioactive substances that may be used as imaging agents in accordance with the embodiments of the disclosure include, but are not limited to, 18 F, 32 P, 33 P, 45 Ti, 47 Sc, 52 Fe, 59 Fe, 62 Cu, 64 Cu, 67 Cu, 67 Ga, 68 Ga, 77 AS, 86 Y, 90 Y.
  • Paramagnetic ions that may be used as additional imaging agents in accordance with the embodiments of the disclosure include, but are not limited to, ions of transition and lanthanide metals (e.g., metals having atomic numbers of 21-29, 42, 43, 44, or 57-71). These metals include ions of Cr, V, Mn, Fe, Co, Ni, Cu, La, Ce, Pr, Nd, Pm, Sm, Eu, Gd, Tb, Dy, Ho, Er, Tm, Yb and Lu.
  • transition and lanthanide metals e.g., metals having atomic numbers of 21-29, 42, 43, 44, or 57-71.
  • These metals include ions of Cr, V, Mn, Fe, Co, Ni, Cu, La, Ce, Pr, Nd, Pm, Sm, Eu, Gd, Tb, Dy, Ho, Er, Tm, Yb and Lu.
  • the imaging agent is a radioactive metal or paramagnetic ion
  • the agent may be reacted with another long-tailed reagent having a long tail with one or more chelating groups attached to the long tail for binding to these ions.
  • the long tail may be a polymer such as a polylysine, polysaccharide, or other derivatized or derivatizable chain having pendant groups to which the metals or ions may be added for binding.
  • chelating groups examples include, but are not limited to, ethylenediaminetetraacetic acid (EDTA), diethylenetriaminepentaacetic acid (DTP A), DOTA, NOTA, NETA, TETA, porphyrins, polyamines, crown ethers, bis-thiosemicarbazones, polyoximes, and like groups.
  • dose refers to the amount of active ingredient given to an individual at each administration.
  • the dose will vary depending on a number of factors, including the range of normal doses for a given therapy, frequency of administration; size and tolerance of the individual; severity of the condition; risk of side effects; and the route of administration.
  • dose form refers to the particular format of the pharmaceutical or pharmaceutical composition, and depends on the route of administration.
  • a dosage form can be in a liquid form for nebulization, e.g., for inhalants, in a tablet or liquid, e.g., for oral delivery, or a saline solution, e.g., for injection.
  • administering means oral administration, administration as a suppository, topical contact, intravenous, intraperitoneal, intramuscular, intralesional, intrathecal, intranasal or subcutaneous administration, or the implantation of a slow-release device, e.g., a mini -osmotic pump, to a subject.
  • Administration is by any route, including parenteral and transmucosal (e.g., buccal, sublingual, palatal, gingival, nasal, vaginal, rectal, ortransdermal).
  • Parenteral administration includes, e.g., intravenous, intramuscular, intra-arteriole, intradermal, subcutaneous, intraperitoneal, intraventricular, and intracranial.
  • Other modes of delivery include, but are not limited to, the use of liposomal formulations, intravenous infusion, transdermal patches, etc.
  • co-administer it is meant that a composition described herein is administered at the same time, just prior to, or just after the administration of one or more additional therapies, for example cancer therapies such as chemotherapy, hormonal therapy, radiotherapy, or immunotherapy.
  • the compounds of the invention can be administered alone or can be co-administered to the patient.
  • Co-administration is meant to include simultaneous or sequential administration of the compounds individually or in combination (more than one compound).
  • compositions of the present invention can be delivered by transdermally, by a topical route, formulated as applicator sticks, solutions, suspensions, emulsions, gels, creams, ointments, pastes, jellies, paints, powders, and aerosols.
  • Formulations suitable for oral administration can consist of (a) liquid solutions, such as an effective amount of the antibodies provided herein suspended in diluents, such as water, saline or PEG 400; (b) capsules, sachets or tablets, each containing a predetermined amount of the active ingredient, as liquids, solids, granules or gelatin; (c) suspensions in an appropriate liquid; and (d) suitable emulsions.
  • Tablet forms can include one or more of lactose, sucrose, mannitol, sorbitol, calcium phosphates, com starch, potato starch, microcrystalline cellulose, gelatin, colloidal silicon dioxide, talc, magnesium stearate, stearic acid, and other excipients, colorants, fillers, binders, diluents, buffering agents, moistening agents, preservatives, flavoring agents, dyes, disintegrating agents, and pharmaceutically compatible carriers.
  • Lozenge forms can comprise the active ingredient in a flavor, e.g., sucrose, as well as pastilles comprising the active ingredient in an inert base, such as gelatin and glycerin or sucrose and acacia emulsions, gels, and the like containing, in addition to the active ingredient, carriers known in the art.
  • a flavor e.g., sucrose
  • an inert base such as gelatin and glycerin or sucrose and acacia emulsions, gels, and the like containing, in addition to the active ingredient, carriers known in the art.
  • compositions can also include large, slowly metabolized macromolecules such as proteins, polysaccharides such as chitosan, polylactic acids, polyglycolic acids and copolymers (such as latex functionalized sepharose (TM), agarose, cellulose, and the like), polymeric amino acids, amino acid copolymers, and lipid aggregates (such as oil droplets or liposomes). Additionally, these carriers can function as immunostimulating agents (i.e., adjuvants).
  • adjuvant refers to a compound that when administered in conjunction with the compositions provided herein including embodiments thereof, augments the composition’s immune response. Generally, adjuvants are non-toxic, have high-purity, are degradable, and are stable.
  • Adjuvants can augment an immune response by several mechanisms including lymphocyte recruitment, stimulation of B and/or T cells, and stimulation of macrophages.
  • the adjuvant increases the titer of induced antibodies and/or the binding affinity of induced antibodies relative to the situation if the immunogen were used alone.
  • a variety of adjuvants can be used in combination with the agents provided herein including embodiments thereof, to elicit an immune response.
  • Preferred adjuvants augment the intrinsic response to an immunogen without causing conformational changes in the immunogen that affect the qualitative form of the response.
  • Preferred adjuvants include aluminum hydroxide and aluminum phosphate, 3 De-O-acylated monophosphoryl lipid A (MPLTM) see GB 2220211 (RIBI ImmunoChem Research Inc., Hamilton, Montana, now part of Corixa).
  • StimulonTM QS-21 is a triterpene glycoside or saponin isolated from the bark of the Quillaja Saponaria Molina tree found in South America (see Kensil et al., in Vaccine Design: The Subunit and Adjuvant Approach (eds. Powell & Newman, Plenum Press, NY, 1995); US Patent No. 5,057,540), (Aquila BioPharmaceuticals, Framingham, MA).
  • adjuvants are oil in water emulsions (such as squalene or peanut oil), optionally in combination with immune stimulants, such as monophosphoryl lipid A (see Stoute etal.,N. Engl. J. Med. 336, 86-91 (1997)), pluronic polymers, and killed mycobacteria.
  • immune stimulants such as monophosphoryl lipid A (see Stoute etal.,N. Engl. J. Med. 336, 86-91 (1997)), pluronic polymers, and killed mycobacteria.
  • Another adjuvant is CpG (WO 98/40100).
  • Adjuvants can be administered as a component of a therapeutic composition with an active agent or can be administered separately, before, concurrently with, or after administration of the therapeutic agent.
  • adjuvants contemplated for the invention are saponin adjuvants, such as StimulonTM (QS-21, Aquila, Framingham, MA) or particles generated therefrom such as ISCOMs (immunostimulating complexes) and ISCOMATRIX.
  • saponin adjuvants such as StimulonTM (QS-21, Aquila, Framingham, MA) or particles generated therefrom such as ISCOMs (immunostimulating complexes) and ISCOMATRIX.
  • Other adjuvants include RC-529, GM-CSF and Complete Freund's Adjuvant (CFA) and Incomplete Freund's Adjuvant (IF A).
  • adjuvants include cytokines, such as interleukins (e.g., IL-1 a and P peptides, IL-2, IL-4, IL-6, IL-12, IL-13, and IL-15), macrophage colony stimulating factor (M-CSF), granulocyte-macrophage colony stimulating factor (GM-CSF), tumor necrosis factor (TNF), chemokines, such as MIPla and and RANTES.
  • M-CSF macrophage colony stimulating factor
  • GM-CSF granulocyte-macrophage colony stimulating factor
  • TNF tumor necrosis factor
  • chemokines such as MIPla and RANTES.
  • Another class of adjuvants is glycolipid analogues including N-glycosylamides, N-glycosylureas and N-glycosylcarbamates, each of which is substituted in the sugar residue by an amino acid, as immuno-modulators or adjuvants (see US Pat. No.
  • Suitable formulations for rectal administration include, for example, suppositories, which consist of the packaged nucleic acid with a suppository base.
  • Suitable suppository bases include natural or synthetic triglycerides or paraffin hydrocarbons.
  • gelatin rectal capsules which consist of a combination of the compound of choice with a base, including, for example, liquid triglycerides, polyethylene glycols, and paraffin hydrocarbons.
  • Formulations suitable for parenteral administration include aqueous and non-aqueous, isotonic sterile injection solutions, which can contain antioxidants, buffers, bacteriostats, and solutes that render the formulation isotonic with the blood of the intended recipient, and aqueous and non-aqueous sterile suspensions that can include suspending agents, solubilizers, thickening agents, stabilizers, and preservatives.
  • compositions can be administered, for example, by intravenous infusion, orally, topically, intraperitoneally, intravesically or intrathecally.
  • Parenteral administration, oral administration, and intravenous administration are the preferred methods of administration.
  • the formulations of compounds can be presented in unit-dose or multi-dose sealed containers, such as ampules and vials.
  • Injection solutions and suspensions can be prepared from sterile powders, granules, and tablets of the kind previously described.
  • Cells transduced by nucleic acids for ex vivo therapy can also be administered intravenously or parenterally as described above.
  • the pharmaceutical preparation is preferably in unit dosage form.
  • the preparation is subdivided into unit doses containing appropriate quantities of the active component.
  • the unit dosage form can be a packaged preparation, the package containing discrete quantities of preparation, such as packeted tablets, capsules, and powders in vials or ampoules.
  • the unit dosage form can be a capsule, tablet, cachet, or lozenge itself, or it can be the appropriate number of any of these in packaged form.
  • the composition can, if desired, also contain other compatible therapeutic agents.
  • the combined administration contemplates co-administration, using separate formulations or a single pharmaceutical formulation, and consecutive administration in either order, wherein preferably there is a time period while both (or all) active agents simultaneously exert their biological activities.
  • Effective doses of the compositions provided herein vary depending upon many different factors, including means of administration, target site, physiological state of the patient, whether the patient is human or an animal, other medications administered, and whether treatment is prophylactic or therapeutic. However, a person of ordinary skill in the art would immediately recognize appropriate and/or equivalent doses looking at dosages of approved compositions for treating and preventing cancer for guidance.
  • the term “pharmaceutically acceptable” is used synonymously with “physiologically acceptable” and “pharmacologically acceptable”.
  • a pharmaceutical composition will generally comprise agents for buffering and preservation in storage, and can include buffers and carriers for appropriate delivery, depending on the route of administration.
  • the terms “pharmaceutically acceptable” or “pharmacologically acceptable” refer to a material which is not biologically or otherwise undesirable, i.e., the material may be administered to an individual in a formulation or composition without causing any unacceptable biological effects or interacting in a deleterious manner with any of the components of the composition in which it is contained.
  • “Pharmaceutically acceptable excipient” and “pharmaceutically acceptable carrier” refer to a substance that aids the administration of an active agent to and absorption by a subject and can be included in the compositions of the present invention without causing a significant adverse toxicological effect on the patient.
  • Non-limiting examples of pharmaceutically acceptable excipients include water, NaCl, normal saline solutions, lactated Ringer’s, normal sucrose, normal glucose, binders, fillers, disintegrants, lubricants, coatings, sweeteners, flavors, salt solutions (such as Ringer's solution), alcohols, oils, gelatins, carbohydrates such as lactose, amylose or starch, fatty acid esters, hydroxymethycellulose, polyvinyl pyrrolidine, and colors, and the like.
  • Such preparations can be sterilized and, if desired, mixed with auxiliary agents such as lubricants, preservatives, stabilizers, wetting agents, emulsifiers, salts for influencing osmotic pressure, buffers, coloring, and/or aromatic substances, and the like., that do not deleteriously react with the compounds of the invention.
  • auxiliary agents such as lubricants, preservatives, stabilizers, wetting agents, emulsifiers, salts for influencing osmotic pressure, buffers, coloring, and/or aromatic substances, and the like.
  • pharmaceutically acceptable salt refers to salts derived from a variety of organic and inorganic counter ions well known in the art and include, by way of example only, sodium, potassium, calcium, magnesium, ammonium, tetraalkylammonium, and the like; and when the molecule contains a basic functionality, salts of organic or inorganic acids, such as hydrochloride, hydrobromide, tartrate, mesylate, acetate, maleate, oxalate and the like.
  • preparation is intended to include the formulation of the active compound with encapsulating material as a carrier providing a capsule in which the active component with or without other carriers, is surrounded by a carrier, which is thus in association with it.
  • carrier providing a capsule in which the active component with or without other carriers, is surrounded by a carrier, which is thus in association with it.
  • cachets and lozenges are included. Tablets, powders, capsules, pills, cachets, and lozenges can be used as solid dosage forms suitable for oral administration.
  • the pharmaceutical preparation is optionally in unit dosage form.
  • the preparation is subdivided into unit doses containing appropriate quantities of the active component.
  • the unit dosage form can be a packaged preparation, the package containing discrete quantities of preparation, such as packeted tablets, capsules, and powders in vials or ampoules.
  • the unit dosage form can be a capsule, tablet, cachet, or lozenge itself, or it can be the appropriate number of any of these in packaged form.
  • the unit dosage form can be of a frozen dispersion.
  • compositions of the present invention may additionally include components to provide sustained release and/or comfort.
  • Such components include high molecular weight, anionic mucomimetic polymers, gelling polysaccharides and finely-divided drug carrier substrates. These components are discussed in greater detail in U.S. Pat. Nos. 4,911,920; 5,403,841; 5,212,162; and 4,861,760. The entire contents of these patents are incorporated herein by reference in their entirety for all purposes.
  • the compositions of the present invention can also be delivered as microspheres for slow release in the body.
  • microspheres can be administered via intradermal injection of drug-containing microspheres, which slowly release subcutaneously (see Rao, J. Biomater Sci. Polym. Ed.
  • the formulations of the compositions of the present invention can be delivered by the use of liposomes which fuse with the cellular membrane or are endocytosed, i.e., by employing receptor ligands attached to the liposome, that bind to surface membrane protein receptors of the cell resulting in endocytosis.
  • compositions of the present invention can focus the delivery of the compositions of the present invention into the target cells in vivo.
  • the compositions of the present invention can also be delivered as nanoparticles.
  • compositions comprising or expressing T cell epitopes, T cell epitope-containing peptides, and T cell epitope -containing proteins associated with binding to a subset of the naturally occurring MHC Class II and/or MHC Class I molecules within the human population.
  • Compositions comprising or expressing one or more of the disclosed peptides (e.g., the amino acid sequences set forth in any one of Tables 1-20) or polynucleotides encoding the same, covering different HLA Class II and/or MHC Class I alleles, capable of generating a treatment acting broadly on a population level are disclosed herein.
  • Such a product should comprise as a first requirement an expression or inclusion of combination of epitopes or peptides that are able to bind the worldwide MHC Class I and/or MHC Class II allele repertoire, and the resulting peptide-MHC complexes should as a second requirement be recognized by the T cells of the subject so as to induce the desired immunological reactions.
  • this is achieved by selecting one or more immunodominant and/or immunoprevalent proteins (e.g., aB. pertussis protein) or subsequences, portions, homologues, variants or derivatives thereof for use in the methods and compositions of the present disclosure, wherein said immunodominant and/or immunoprevalent proteins or subsequences, portions, homologues, variants or derivatives thereof comprise two or more epitopes that are immunodominant and/or immunoprevalant.
  • immunodominant and/or immunoprevalent proteins e.g., aB. pertussis protein
  • An additional object of the invention is to provide proteins, peptides, or nucleic acids containing or expressing epitopes or combinations of such proteins, peptides or nucleic acids which have a sufficient solubility profile for being formulated in a pharmaceutical product, preferably which have acceptable estimated in vivo stability.
  • One further objective of the invention is to select epitopes for use in the compositions and methods described herein, based on one or both of their immunodominance or immunoprevalence.
  • a still further object of the invention is to select such epitopes and epitopes combinations not only in accordance with those embodiments previously described, but also those epitopes and epitope combinations capable of eliciting a B cell response and T cell response (e.g., selecting one or more peptides for use in the methods and compositions described herein capable of generating a T cell and antibody response in a subject).
  • kits for modulating, eliciting, or detecting T cells responsive to one or more Bordetella peptides or proteins.
  • proteins and peptides described herein comprise, consist of, or consist essentially of: one or more amino acid sequences selected from any one of those sequences set forth in Tables 1-20, or a subsequence, portion, homologue, variant or derivative thereof; a fusion protein comprising one or more amino acid sequences selected from any one of those sequences set forth in Tables 1-20; a pool of 2 or more peptides selected from the amino acid sequences set forth in any one of Tables 1-20, or a polynucleotide that encodes one or more peptides or proteins, comprising, consisting of, or consisting essentially of an amino acid sequence selected from any one of those sequences set forth in Tables 1-20, or a subsequence, portion, homologue, variant or derivative thereof.
  • the Bordetella is one or more of B. pertussis or a variant thereof. Further description and embodiments of such methods and compositions are provided in the definitions provided herein, and a person skilled in the art will recognize that the methods and compositions can be embodied in numerous variations, changes, and substitutions or as may occur to or be understood by one skilled in the art without departing from the invention.
  • the words “comprising” (and any form of comprising, such as “comprise” and “comprises”), “having” (and any form of having, such as “have” and “has”), “including” (and any form of including, such as “includes” and “include”) or “containing” (and any form of containing, such as “contains” and “contain”) are inclusive or open-ended and do not exclude additional, unrecited elements or method steps.
  • “comprising” may be replaced with “consisting essentially of’ or “consisting of’.
  • the phrase “consisting essentially of’ requires the specified integer(s) or steps as well as those that do not materially affect the character or function of the claimed invention.
  • the term “consisting” is used to indicate the presence of the recited integer (e.g., a feature, an element, a characteristic, a property, a method/process step or a limitation) or group of integers (e.g., feature(s), element(s), characteristic(s), propertie(s), method/process steps or limitation(s)) only.
  • words of approximation such as, without limitation, “about”, “substantial” or “substantially” refers to a condition that when so modified is understood to not necessarily be absolute or perfect but would be considered close enough to those of ordinary skill in the art to warrant designating the condition as being present.
  • the extent to which the description may vary will depend on how great a change can be instituted and still have one of ordinary skilled in the art recognize the modified feature as still having the required characteristics and capabilities of the unmodified feature.
  • a numerical value herein that is modified by a word of approximation such as “about” may vary from the stated value by at least ⁇ 1, 2, 3, 4, 5, 6, 7, 10, 12 or 15%.
  • compositions and/or methods disclosed and claimed herein can be made and executed without undue experimentation in light of the present disclosure. While the compositions and methods of this invention have been described in terms of preferred embodiments, it will be apparent to those of skill in the art that variations may be applied to the compositions and/or methods and in the steps or in the sequence of steps of the method described herein without departing from the concept, spirit and scope of the invention. All such similar substitutes and modifications apparent to those skilled in the art are deemed to be within the spirit, scope and concept of the invention as defined by the appended claims.
  • each dependent claim can depend both from the independent claim and from each of the prior dependent claims for each and every claim so long as the prior claim provides a proper antecedent basis for a claim term or element.

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Abstract

La présente invention concerne des compositions, comprenant des mégapools d'épitope, et des méthodes de détection de la présence : d'une Bordetella ou d'une réponse immunitaire pertinente pour une infection par Bordetella notamment des cellules sensibles à un ou plusieurs peptides ou protéines de Bordetella comprenant, consistant en, ou consistant essentiellement en : une ou plusieurs séquences d'acides aminés, des protéines de fusion, un pool d'au moins 2 peptides, ou des polynucléotides qui expriment les séquences d'acides aminés sélectionnées parmi celles présentées dans l'une quelconque des tables 1-20 (NO ID SÉQ ID : 1 à 2598). L'invention concerne en outre des vaccins, des diagnostics, des thérapies et des kits, contenant ces protéines ou peptides.
PCT/US2023/069274 2022-06-28 2023-06-28 Épitopes de lymphocytes t de bordetella, mégapools et utilisations associées WO2024006842A2 (fr)

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